HPMTLE REVIEW NOTES

MTLE BOARD EXAM TABLE OF SPECIFICATION (TOS) Reduced oxygen Cell injury
Histopathologic techniques, Med. Tech Laws, Related Laws supply; chemical injury;
& Code of Ethics microbial infection
Acute and transient Acute reversible injury
Histopathologic Techniques: 85% Cellular swelling fatty
Tissue processing: 40% changes
Routine: 30% Irreversible injury -> cell
Special: 10% death
Cytology: 10% Necrosis
Staining: 20% Apoptosis
Routine: 15% Metabolic alterations, Intracellular
Special: 5% genetic or acquired ; accumulations;
Autopsy: 5% chronic injury calcification
Special procedures: 5% Cumulative. Sublethal Cellular aging
Clerical/ Logging: 5% injury over long life
span
Medtech Laws, Related Laws and Code of Ethics: 15%
Med. Tech Laws: 10% These changes include:
Related Laws: 3%  Hypertrophy – increased cell mass
Code of Ethics: 2%  Atrophy – decreased cell mass
 Hypoplasia- fewer cells than what is deemed a normal amount
(usually benign) e.g. micromastia (post pubertal female breast
underdevelopment)
PATHOLOGY  Hyperplasia – increased cell number. Under control of a normal
is the study of the structural and functional causes of human proliferation regulatory mechanism. A result of external stimuli
disease. The four aspects of a disease process that form the e.g. callus formation on skin exposed to trauma/pressure.
 Neoplasia- similar to hyperplasia but denotes abnormal
core of pathology are. multiplication due to loss of normal proliferation regulation and
absence of stimuli. Also known as “cells manifesting
 Etiology – cause of the disease. hyperplasia with atypia”.
 Pathogenesis – mechanism(s) of disease  Dysplasia- a change in the normal shape, size and
organization, usually a response to chronic irritation e.g.
development. cigarette smoking or inflammation. Change is reversible if
 Morphologic change – the structural alterations stimulus is removed otherwise the cells will become neoplastic.
induced in cells and tissues by the disease.  Metaplasia – change from one mature cell type to another.
Change are also reversible if stimulus is removed, otherwise
 Clinical Significance – the functional cells become anaplastic.
consequences of the morphologic changes.  Anaplasia – a reversal in differentiation (dedifferentiation) or
 Adaptation –occurs when physiologic or loss of structural and functional differentiation of normal cells.
pathologic stressors induce a new state that These cells changes are not reversible in nature.
Characteristics of cancerous tumours.
changes the cell but otherwise preserves its
 Reversible injury – denotes pathologic cell changes that can be
viability in the face of the exogenous stimuli. restored to normalcy if the stimulus is removed or if the cause
is mild.
Cellular Response to Injury  Irreversible injury - occurs when stressors exceed the capacity
of the cell to adapt (beyond a point of no return) and denotes
Nature of injurious Cellular Response permanent pathologic changes that cause cell death.
Stimulus
Altered physiological Cellular adaptations Cell death occurs primarily through 2 morphologic and
stimuli; some mechanistic patterns denoted necrosis and apoptosis.
nonlethal, injurious
stimuli Features of Necrosis and Apoptosis
Increased demand, Hyperplasia, hypertrophy Feature Necrosis Apoptosis
increased stimulations Cell size Enlarged (swelling) Reduced
(e.g. by growth factors, (shrinkage)
hormones) Nucleus Pyknosis- Fragmentation into
Decreased nutrients, Atrophy >karyorrhexis- nucleosome-size
decreased stimulation >karyolysis fragments
Chronic irritation Metaplasia Plasma Disrupted Intact; altered
membrane structure,
(physical or chemical) especially
orientation of lipids

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 HPMTLE REVIEW NOTES

Cellular contents Enzymatic Intact; may be 3. Gangrenous necrosis – is not a specific pattern but
digestion; may leak released in is rather just coagulative necrosis as applied to
out of the cells apoptotic bodies ischemic limbs (wet gangrene).
Adjacent Frequent None 4. Caseous necrosis – tuberculosis lesions, appears
inflammation as soft, friable, cheesy material and
Physiologic or Invariable Other physiologic,
microscopically as amorphous eosinophilic
pathologic role pathologic Means of
(culmination of eliminating material with cell debris.
irreversible cell unwanted cells, 5. Fat necrosis – is seen in adipose tissues, lipase
injury) may be pathologic activation (e.g. injured pancreatic cells or
after some forms of macrophages. Grossly these are white, chalky
cell injury, areas (fat saponification), histologically there are
especially DNA vague cell outlines and calcium deposition.
damage. 6. Fibrinoid necrosis – is a pathologic pattern
resulting from antigen-antibody (immune complex)
 Necrosis – more common type of cell death, involving severe deposition in blood vessels. Microscopically there
cell swelling, denaturation and coagulation of proteins,
breakdown of cellular organelles, and cell rupture. are bright pink amorphous material (protein
 Apoptosis – programmed cell death deposition) in arterial walls often associated
 Autophagy – is an adaptive response of cells to nutrient inflammation and thrombosis.
deprivation; it is essentially a self cannibalization to maintain
viability (e.g. degenerative disorders of muscle and nervous
system).
Mechanisms of cell injury
 Pyknosis- irreversible condensation of chromatin in the nucleus 1. Depletion of ATP
of a cell undergoing necrosis or apoptosis (small, dense 2. Mitochondrial Damage
nucleus) 3. Influx of Calcium and loss of calcium homeostasis
 Karyorrhexis – is the destructive fragmentation of the nucleus 4. Accumulation of oxygen derived free radicals
of a dying cell whereby its chromatin is distributed irregularly
throughout the cytoplasm (fragmented nucleus) (oxidative stress)
 Karyolysis – complete dissolution of the chromatin of a dying 5. Defects of membrane permeability
cell due to the enzymatic degradation of endonucleases (faint, 6. Damage to DNA and proteins
dissolved nucleus).
Overview of Inflammation
Causes of cell injury
1. Oxygen deprivation (hypoxia) – ischemia (loss of The response of vascularized living tissue to injury. It may
blood supply), inadequate oxygenation (e.g. be evoked by microbial infection, physical agents, chemical,
cardiorespiratory failure) and loss of oxygen- necrotic tissue, or immune reactions.
carrying capacity of blood (e.g. anemia, carbon
monoxide poisoning) Two main components: vascular wall response and
2. Physical agents – trauma, heat, cold, radiation and inflammatory cell response.
electric shock
3. Chemical agents and drugs – therapeutic drugs, Onset Duration Cells
poison, environmental pollutants and social stimuli involved
(alcohol and narcotics) Acute Early Short (minute PMNs
4. Infections agents – viruses, bacteria, fungi, and inflammation or days)
parasites involving fluid
5. Immulologic reactions – autoimmune diseases exudation
6. Nutritional imbalances/ excesses (edema)
Chronic Later Longer Lymphocytes
General tissue patterns of necrosis:
inflammation (weeks or and
1. Coagulation necrosis – the most common pattern,
months) w/ Macrophages
predominated by protein denaturation with
blood vessel
preservation of the cell and tissue framework. All
proliferation
tissues except the brain.
and fibrosis
2. Liquefaction necrosis – occurs when autolysis or
heterolysis predominates over protein
denaturation (localized bacterial infections or
abscess).

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 HPMTLE REVIEW NOTES

Five cardinal signs of Inflammation Cases referred to NBI or PNP

• Accident
1. Calor (warmth): due to vascular dilation • Suicide, homicide, poisoning, burns, criminal
2. Rubor (erythema): due to vascular dilation and violence, trauma, unusual circumstances
coagulation • Death w/o medical attendance
3. Tumor (edema): due to increased vascular • DOA (Dead on Arrival)
permeability • Abortion, death under anesthesia or surgery,
4. Dolor (pain): due to mediator release alcoholism
5. Functio laesa (loss of function): due to pain,
edema, tissue injury, and/or scar Techniques:
1. Virchow
 Edema – excess fluid in interstitial tissue or body cavities and  Organs removed one by one (organ by organ)
can either an exudate or a transudate.
 Exudate – is an inflammatory, extravascular fluid with cellular
 First the cranial cavity is exposed and from the
debris and high protein concentration (specific gravity 1020 or back, the spinal cord, followed by the thoracic,
more). cervical and abdominal organs.
 Transudate – is excess, extravascular fluid with low protein  Most widely used method.
content (specific gravity of 1.012 or less), ultrafiltrate of blood
plasma resulting from elevated fluid pressures of diminished  Advantage: each organ can be studied in detail.
osmotic forces. However the anatomicopathologic relationships of
 Pus – purulent inflammatory exudate rich in neutrophils and organs are not preserved thus cannot be studied.
cell debris.
2. Rokitansky
AUTOPSY  In situ (in the original place) dissection combined
Purpose: To ascertain cause of death with en bloc (all together) removal
Causes of Death:  Dissection starts in the neck and trials down, and
1. Homicidal the organ is removed as a bloc as well.
2. Suicidal  This technique involves the in situ dissection in
3. Accidental part combined with en bloc removal.
4. Natural causes
 This technique is commonly preferred whenever
the pathologist wants to limit the spread of
Value:
infection such as HIV, Hepatitis B, etc.
• Provides opportunity of studying nature of disease
 Disadvantage: the organs cannot be studied in
• See results of certain treatment
detail.
• Reveals congenital & acquired abnormalities
• To visualize normal & abnormal parts of the body
3. Ghon
• Elimination of fear
• Recognition of familial or hereditary disorders  Removed as organ blocks.
• Provides reliable source of vital statistics  En bloc removal of viscera into thoracic, intestines,
• Aids in the study of epidemic diseases upper abdominal, lower abdominal, brain and
neck.
Who authorizes autopsy?  In this method the thoracic, cervical, abdominal
• Next of kin & 2 witnesses organs and the uro genital system are removed as
Next of Kin? organ blocks.
• spouse
• children of legal age 4. Letulle
• father  In this method thoracic, cervical, abdominal and
• mother pelvic organs are removed en mass but
• next nearest relative subsequently dissected as organ block.
 En masse removal then subsequently dissected
Pediatrics, stillbirths, prematures? into organ blocks
• father of legal age or “emancipated  Best for routine inspection & preservation of
minor” connections between organ and organ systems.
• mother if of legal age without husband or dead  The organ blocks can be studied in details.
• husband or minor husband  Fast, awkward to handle
• maternal grandparent (grandfather) if father is
minor, not supporting wife, or mother is minor

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 HPMTLE REVIEW NOTES

Adult Autopsies
• External description Restricted Autopsies
• Radiologic examination • Restriction of skin incision
• Y shaped incision • Autopsies thru surgical wounds
• Removal of material from abdomen for • Autopsies thru anus & vagina
microbiology • Needle autopsies (liver, heart, lungs, kidneys)
• Collection of effusions & exudates
• Record diaphragm height & level of liver edge Special Problems in Autopsy
• Search for hernias • Adhesions
• Incision of anterior abdominal musculature & • Hernias
breasts • Lesions on face, arms, hands
• Search for pneumothorax need special permission
• Cut lower ribs • Pneumothorax, pneumomediastinum, subcuta-
• Removal of chest plate neous emphysema, free air in abdomen
• Removal of thymus fat pad
• Collection of pericardial contents Estimation of Time of Death
• Collection of blood from heart or peripheral system 1. Rigor mortis
for micro & biochemical studies • Normal stiffening occurs in 2-3 hours after death
• ID of carotid, subclavian, femoral arteries for • Complete in 6-8 hours
embalmer • Persists for 12-36 hours
Pediatric Autopsies • Begins with small muscle groups
• Potter & Craig (lungs, liver, kidney, thymus, brain • Delayed in cold, emaciation, extreme obesity
– minimal requirement) • Hasten with heat, strenuous exercise,
• External examination, check for malformations convulsions, & in infants
• Examine placenta & umbilical cord 2. Livor mortis
• Skull opened by Beneke technique • Post mortem lividity
• Open chest cavity under water to show • Sinking of blood in dependent parts
pneumothorax • Fixed in 6-8 hours
• En masse removal is preferred • Completed in 8-12 hours
Post Operative Autopsies • Determine whether body position is changed
• Possible medico legal implications 3. Algor mortis
• Most experienced pathologist • Body cooling after death
• Surgeon should be present • Postmortem intervals:
• Incision not carried out thru operative wounds 98.6°F (rectal) per 1.5 hours
• Document with photographs 37°C (rectal) per 0.83 hour
• Dictate descriptions, do not interpret Chemical means:
Immediate Autopsy Program • Chemical determinations in serum & CSF
• Tissues obtained trimmed. after somatic death • Aminonitrogen, NH3, inorganic phosphates
• Investigation of pathophysiologic effects of shock, Stomach contents:
trauma, & metabolic diseases made possible • Stomach empties 2-3 hours after meal, delayed by
• Feasible to apply EM, histochem., analytical psychologic factors or physical trauma (esp. head)
biochem. Putrefaction:
• Extremely difficult
2 Phases: • Factors: bacteria, moisture, temperature of
1. Rapid Sampling Phase environment, medium where body lies
• Done within minutes of death pronouncement • Hastened in warm environment
• Samples from: heart, lung, liver, kidneys, • Rapid in air than water
adrenals, pancreas, skeletal muscles, aorta, • Rapid in water than when buried
colon, prostate, breast in females • Slow in NB & extreme cold or dryness
• Team of 6
• Fixatives: 4% phosphate buffered formalin & 1% FRESH TISSUE EXAMINATION
glutaraldehyde
• Blood sample collected for chemistry Methods if Fresh Tissue Examination

2. Routine Phase: 1. Teasing or dissociation – process whereby a selected

• Uses standard dissection, fixation, & sampling tissue specimen is immersed in a watch glass containing

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 HPMTLE REVIEW NOTES

isotonic salt solution, carefully dissected or separated, and 4. Frozen section – this method is normally utilized when a
examined under bright field microscope, or stained with rapid diagnosis of the tissue in question is required and is
differential dyes. especially recommended when lipids and nervous tissue
elements are to be demonstrated. Very thin slices – 10-15
2. Squash preparation (crushing) – is a process whereby um in thickness are cut from a fresh tissue frozen on a
small pieces of tissue <1mm are placed in a microscopic microtome with carbon dioxide or on a cryostat, a cold
slide and forcibly compressed with another glass slide or chamber kept at an atmospheric temperature of -10C to
with a cover glass. If necessary, a vital stain may be placed -20C. The frozen sections are then transferred to a slide, and
at the junction of the slide and the cover glass, and allowed processed for light microscope study. The majority of non-
to be absorbed by the tissue through capillary attraction. fatty unfixed tissues are sectioned well at temperature -10C
and -25C.
3. Smear preparation – is the process of examining
sections or sediments, whereby cellular materials are spread Applications of frozen sections in histopathology
over a slide by means of a wire loop or applicator, or by
making an apposition smear with another slide. This 1. Rapid pathologic diagnosis during surgery
technique is especially useful in cytological examinations, 2. Diagnostic and research enzyme histochemistry
particularly for cancer diagnosis. 3. Diagnostic and research demonstration of soluble
a. Streaking – with an applicator stick or a platinum loop, the substances such as lipids and carbohydrates
material is rapidly and gently applied in a direct or zigzag line 4. Immunofluorescent and immunohistochemical
throughout the slide, attempting to obtain a relatively uniform staining
distribution of secretion. To thin and too thick smears have 5. Some specialized silver stains, particularly in
to be avoided, since they make the tissues unsuitable for neuropathology
examination.
Most common methods of freezing
b. Spreading- a selected portion of the material is transferred
to a clean slide and gently spread into a moderately thick film 1. Liquid Nitrogen – most rapid and commonly
by teasing the mucous strands apart with an applicator stick. available freezing agents.
This method is a little more tedious than streaking, but Main disadvantage: soft tissue is liable to crack
has the advantage of maintaining cellular interrelationships due to rapid expansion of ice within the tissue
of the material to be examined. It is especially recommended producing ice crystal and freeze artifacts.
for smear preparations of fresh sputum and bronchial Remedy: freeze the tissue with Isopentane, OCT,
aspirates, and also thick mucoid secretions. or Freon 2.2 (high thermal conductivity).
2. Isopentane cooled by liquid nitrogen
c. Pull –Apart – is done by placing a drop of secretion or 3. Carbon dioxide gas
sediment upon one slide and facing it to another clean slide. 4. Aerosol sprays
The material disperse evenly over the surface of the two
slide. Slight movement of the two slides in opposite
directions may be necessary to initiate the flow of materials. ROUTINE TISSUE PROCESSING
The two slides are then pulled apart with a single
uninterrupted motion and the specimen placed under the A. FIXATION
microscope for immediate examinations or applied with vital process of preserving cells and tissue constituents in a
stain. Useful is useful for preparing smears of thick condition identical to that existing during life.
secretions such as serous fluids, concentrated sputum, To maintain adequate fixation – 4-6 hrs, size: 2 cm2 and no
enzymatic lavage samples from the gastrointestinal tract and more than 4mm thick.
blood smear.
Two Important Goals of Fixation
d. Touch preparation (Impression smear) – is a special 1. Primary Goal: Preserve the morphologic and chemical
method of smear preparation whereby the surface of a integrity of the cell
freshly cut piece if tissue to brought into contact and pressed 2. Secondary Goal: Harden and preserve the tissue for
on to the surface of a clean glass slide, allowing the cells further handling
to be transferred directly to the slide for examination by
phase contrast microscope or stained for light microscope Most important reaction for maintaining tissue appearance:
study. It has an advantage in that the cells may be examined Stabilization of proteins
without destroying their actual intercellular relationship and Two mechanisms of Action
without separating them from their normal surroundings.  Additive
 Non Additive

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 HPMTLE REVIEW NOTES

Factors involved in fixation Aldehyde fixatives

1. pH (Hydrogen Ion Concentration) –satisfactory pH 6 and Formaldehyde
8 10% Formol saline –recommended for CNS
2. Temperature – heat for bacteriology, room temperature tissues and general post mortem tissues for histo
(25C traditional) - Mast cells. 40C – most lab, electron 10% Neutral buffered formaldehyde –
microscopy (0 - 4C). Formalin heated 60C – rapid/ urgent preservation and storage of surgical, post mortem
biopsy specimen. Formalin 100C – tissues with TB and research specimens.
3. Thickness - 1-2 mm2 for electron microscopy, 2cm2 for Formol Corrosive – recommended for routine post
light microscopy or thin (no more than 0.4cm for light mortem tissues
microscopy) Alcoholic formalin (Gendre’s fixative) -
4. Osmolality – slightly hypertonic solution (400-500 mOsm, Glutaraldehyde
isotonic solutions 340 mOsm) Paraformaldehyde
5. Concentration – 10% formalin, 3% glutaraldehyde 0.25%
glutaraldehyde – ideal concentration for immunoelectron Metallic fixative
microscopy. Mercuric Chloride fixatives– most common metallic fixative
6. Duration of fixation – Buffered formalin – 2-6 hrs Zenkers
Prolong fixation: inhibit enzyme activity and immunological Zenker-formol (Helly’s solution) – microanatomical
reactions, cell shrinkage and hardening of tissues. fixative for pituitary gland, bone marrow and blood
containing organs such as liver and kidney.
Electron microscopy - 3 hrs then places in a holding buffer Heidenhans –Susa –tumor biopsies of the skin
B-5 fixative – bone marrow biopsies
Practical Consideration Chromate fixatives
1. Speed – ASAP to prevent autolysis and putrefaction Chromic acid
2. Rate of penetration - ~1mm/hr then slows down as the Potassium Dichromate
tissue goes deeper Regard’s Fluid (Muller’s)
3. Volume – 10-25x of the volume of tissue to be fixed. ~20x Orth’s Fluid
the volume of the tissue – maximum effectiveness. Lead fixatives
4. Duration- Picric acid fixatives
Fibrous organs: Uterus or GIT (need longer) Bouin’s – embryos and pituitary biopsies, 6-24 hrs
Loosely textured tissues (Biopsies and Scrappings). Can be Brasil’s Alcoholic picroformol -
cut down by heat, vacuum, agitation or microwave
Brain – porous, Bone marrow – refrigeration (mitosis up to Glacial acetic acid
30 minutes) Usually incorporated in compound fixatives
Solidifies at 17°C
Fixative types according to composition Precipitates nucleoproteins, chromatin materials
 Simple – made up of only one component Not for cytoplasmic fixation
substance
 Compound – made up of two or more fixatives Alcoholic fixative
which have been added together to obtain the Methanol (100%)
optimal combined effect if their individual actions Isopropyl alcohol (95%)v- touch preparations,
upon the cells and tissue constituent. Wrights- Giemsa
Ethanol (70% - 100%) 18-24 hrs
Fixative types according to action Carnoy’s fixative – brain tissues (rabies),
1. Microanatomical – those that permit the general chromosomes, lymph glands and urgent biopsies,
microscopic study of tissue structures without altering the 1-3 hrs
structural pattern and normal intracellular relationship of Newcomer’s – mucopolysaccharides and nuclear
tissue in question. proteins, 12-18 hrs
2. Cytological – those preserve specific parts and particular
microscopic elements of the cell itself. Osmium tetroxide
Nuclear – with glacial acetic acid, pH 4.6 or less Flemming’s
Cytoplasmic – never glacial acetic acid, pH of 4.6 Flemming’s without acetic acid
or more
3. Histochemical – preserves the chemical and constituents Trichloroacetic acid
of cells and tissues. Precipitates protein
Weak decalcifying agent
Has a softening effect on dense tissues

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 HPMTLE REVIEW NOTES

False negative results

Acetone fixatives • Ab is inappropriate, denatured, or used at wrong
Used at ice cold temperature concentration
For diffusible enzymes (phosphatase and lipase) • (+) loss of Ag thru autolysis & or diffusion
For fixing brain tissues: rabies (Negri bodies) • Ag is below detection level

Heat fixation Actin - Contractile protein for motility

Thermal coagulation of tissue proteins • For identification of smooth muscle cells &
For frozen tissue sections and preparations of myoepithelial cells
bacteriologic smears • Used for rhabdomyosarcoma (sarcomeric)
Alkaline Phosphatase
Microwave technique • Membrane bound glycoprotein
 Dipolar molecules, causing oscillation at a • Hepatic, osseous, renal, placental (PLAP)
frequency 2450mHz • Gonadal & extragonadal germ cell tumors
 Special stains, histochemistry, enzyme Alpha-1-antichymotrypsin & Alpha-1-antitrypsin
histochemistry and immuno- cytochemical • An acute-phase plasma protease inhibitor
techniques. • Synthesized in the liver
• Marker for histiocytes and reticulum cells
ENZYME HISTOCHEMISTRY • Limited diagnostic utility
Fixative: 4% formaldehyde or formol saline Alpha Fetoprotein
• Major plasma component of fetus
ELECTRON MICROSCOPY • From liver & visceral endoderm of yolk sac
Fixative: osmium tetroxide, glutaraldehyde and • Yolk sac tumor or Endodermal sinus tumor
paraformaldehyde at 4C Angiotensin- converting Enzyme
• In vascularized endothelium of lungs & other
IMMUNOHISTOCHEMISTRY tissues
Application of immunologic principles & techniques to the • In epithelial cells of renal proximal tubules &
study of cells & tissues intestinal mucosa
Two most common procedures: • Detected in renal cell carcinoma
1. Peroxidase- antiperoxidase immune B72.3
complex method • Consistently expressed by wide variety of
2. Biotin- avidin immunoenzymatic method adenocarcinomas
“antigen unmasking” techniques • Used in the differential diagnosis of mesothelioma
Digestion with proteolytic enzymes, microwave treatment, & adenocarcinoma
combined action of heat and pressure (pressure cooker) BCA 225
• From human breast carcinoma tissue
Advantages: • However, (+) also in carcinomas of kidney, ovary,
• Remarkable sensitivity and specificity & lungs
• Applicability to routinely processed tissue CA 19.9
• Feasibility of an accurate correlation with • Carbohydrate antigen
traditional morphologic parameters • Pancreatic CA, pancreatobiliary CA, & trasitional
• Compatible with most fixatives cell CA
• Can be used in decalcified material or in CA 125
previously stained tissues • Useful in mucinous epithelial ovarian tumors
• Also in cervix, endometrial, GIT, & breast tissues
False positive results Cadherins
• Cross reactivity of Ab with Ag different from being • Located in desmosomes
sought • In CAs presence inversely correlated with
• Non-specific binding of Ab to the tissue differentiation
• Presence of endogenous peroxidase in some CEA
cellular elements • GIT adenocarcinomas (colon)
• Entrapment of normal tissues by tumor cells • Pancreas, lungs, thyroid medullary CA
• Release of soluble proteins from cytoplasm of CD44v6
normal cells invaded by the tumor • Breast CA
CD45 (LCA – Leukocyte Common Antigen)
• Lymphoid tissues

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• Lymphoma Nestin
CD63 • Abundant in neuroepithelial stem cells
• Melanoma • Also seen in neuroectodermal tumors & gliomas
CD99 Neurofilaments
• Ewings sarcoma • Tumors of neuronal origin or neural differentiation
Chomogranin • e.g. Medulloblastoma, neuroblastoma,
• Neuroendocrine tumors retinoblastoma
• Pan-endocrine marker Neuron Specific Enolase (NSE)
Cytokeratin • Neuroectodermal & neuroendocrine neoplasms
• For tumors with epithelial differentiation • Carcinoid tumors & malignant melanoma
• At 20 subclasses Osteonectin
Desmin • Osteosarcoma & other osteoid forming lesions
• Smooth muscle & skeletal muscle tumors p53
Epithelial Membrane Antigen (EMA) • Most common genetic alteration in human tumors
• Primary epithelial occurs here
• Also in mesotheliomas, meningiomas, • Involved in apoptosis & maintenance of genomic
mesenchymal neoplasm stability
Estrogen Receptor Progesterone Receptor
• Requested for breast CA patients for management • Requested for breast CA patients for management
Factor VIII Prostatic Specific Antigen (PSA)
• Endothelial cell differentiation • Prostatic CA
Ferritin S-100
• Iron binding protein • In glial & Schwann cells, melanocytes,
• Liver cell CA & Breast CA chondrocytes, adipocytes, myoepithelial cells
Glial Fibrillary Acidic Protein (GFAP) • Malignant melanoma
• Present in normal, reactive, & neoplastic Serotonin
astrocytes • Neuroendocrine tumors, carcinoid tumor
• In developing, reactive, & neoplastic ependymal Synaptophysin
cells • Expressed in normal reactive cells of
• In developing & neoplastic oligodendrocytes neuroectodermal & neuroendocrine tumors
• Also in peripheral nerve tumors, & in mixed tumors • Pheochromocytoma, thyroid medullary CA,
of the salivary glands and sweat glands endocrine pancreatic tumor, carcinoid tumor
HMB-45 Thyroglobulin
• Melanoma, neuralcrest-derived tumors, • Thyroid differentiation, thyroid neoplasm
angiomyolipomas of kidneys, tuberous-sclerosis Vimentin
complex • Mesenchymal nature
Human Chorionic Gonadotropin • Endothelial cells, fibroblasts, & vascular smooth
• Normally secreted by syncitiotrophoblasts muscle cells
• Trophoblastic differentiation in germ cell tumor
• Ectopic hCG production of other neoplasm DECALCIFICATION
Laminin  done in between fixation and other processes.
• Tumors that produces basement membrane  Bones, teeth and other calcified tissues (e.g.
• e.g.. CA, smooth muscle tumors, tumors of Calcified lung tissues).
endothelial cells  Also known as “Demineralization”
Myelin proteins  Procedure whereby calcium or lime salts are
• Central & peripheral myelin sheaths removed from tissues following fixation (acids – to
• In oligodendrocytes & Schwann cells form soluble calcium salts or chelating agents- to
Myoglobulin bind into calcium ions).
• Specific for striated muscle (skeletal & myocardial)
• For rhabdomyosarcomas Calcium can be removed by any of the following agents:
• Low sensitivity 1. Acids
• Only for well-differentiated tumors 2. Chelating agents
Myosin 3. Ion exchange resins
• 2 types: smooth muscle type (non sarcomeric 4. Electrical ionization (Electrophoresis)
type) skeletal muscle type (sarcomeric)
• For skeletal muscle differentiation in tumors

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 HPMTLE REVIEW NOTES

Acid decalcifying agents POST DECALCIFICATION

Nitric acid –most common and the fastest decalcifying agent Saturated lithium carbonate solution or 5-10% aqueous
(5-10%) sodium bicarbonate solution for several hrs.
5-10% Nitric acid:12-24hrs Running tap water 30 mins: small samples, 1-4 hrs larger
Formol-Nitric acid: 1-3 days specimens
Perenyi’s fluid: 2-7 days Acid decalcified tissues for frozen sections: formol saline
Phloroglucin- Nitric acid: 12-24 hrs containing 15% sucrose or phosphate-buffered saline with
Hydrochloric acid – slower and greater distortion but 12-20% sucrose at 4C before freezing
produced better nuclear staining.
Von Ebner’s Fluid B. DEHYDRATION
Formic acid – moderate acting, better nuclear staining and  A step in tissue processing in which the
less distortion SAFER than nitric acid and HCl. 2-7 days intercellular and extracellular water from the tissue
Formic acid-Sodium Citrate: 3-14 days are removed after fixation and prior to wax
Tricholoracetic acid: 4-8 days infiltration
Sulfurous acid: suitable only to minute piece/s of bone/s  Amount of dehydrating agent: 10x the volume of
Chromic acid (Flemming’s fluid): maybe used as BOTH the specimen or more
fixative and decalcifying agent  EDTA decalcified – X 70% alcohol O water or
Citric acid (pH 4.5): 6 days storing overnight in formol saline or PBS

Chelating agents Dehydrating Agents- remove the fixative and water from the
EDTA: for immunohistochemical stain or enzyme staining, tissue and replace them with dehydrating agent with
and for electron microscopy dehydrating fluid in preparation for impregnation.
Disodium salt (10%) – simply aqueous or buffered
pH 7.0-7.4 Alcohol –most common, recommended, best dehydrating
Tetrasodium solution – alkaline adjust to pH 7.4 agent. 37C- hasten dehydration
using conc. Acetic acid Methyl alcohol – blood and tissue films.
Ion exchange resin – hastens decalcification by removing Butyl alcohol – plant and animal micro-techniques
calcium ions from formic acid containing decalcifying Ethyl alcohol
solutions. X recommended for fluids containing mineral Notes to Remember when Using Alcohol
acids such as nitric acid or HCl. Decalcifying agent is 20-30x 1. Strength of initial alcohol required
2. Length of storage
the volume of the tissue. 1-14 days. 3. Temperature

Electrophoresis (Electrical Ionization): shorten time of ***Anhydrous Copper Sulfate

decalcification because of heat and electrolytic action in the Serves as an indicator that will accelerate and ensure the complete
process. dehydration. Blue – incomplete dehydration

Factors that affect decalcification Acetone – urgent biopsies

Dioxane (Diethlyne Dioxide) – excellent dehydrating and
1. Structure clearing agent (miscible with water, melted paraffin, alcohol
2. Temperature : 18-30C and xylol)
3. Volume of the Solution: 20:1 (fluid: tissue) Cellosolve (Ethylene glycol monoethyl ether)
4. Time: ideal 12-24hrs Triethyl phosphate
Tetrahydrofuran (THF): X eye and skin irritant, conjunctival
Measuring extent of decalcification irritation.
1. Physical and Mechanical test (e.g. pricking, slicing
or bending) C. CLEARING
2. X-ray or radiological method (e.g. FAXITRON or  Also known as decalcoholization
Kodak X-OMAT)  A process of removal of the dehydrating agent and
3. Chemical method (conc. Ammonium Hydroxide- replacement of a fluid (clearing agent) that is
neutralization of acid solution (clear) ; saturated miscible with both the dehydrating agent and
aqueous Ammonium Oxalate-stand for 30 embedding medium
minutes, cloudiness of the solution indicates
incomplete decalcification ) CLEARING AGENTS
1. Xylene- most common, 0.5 to 1 hr tissue block
<5mm
2. Toluene- 1-2 hrs

PATRICK R. DE VERA,RMT 9
 HPMTLE REVIEW NOTES

3. Benzene 3. Plastic embedding rings and base mold

4. Chloroform 4. Disposable embedding mold
5. Cedarwood Oil- 2-3 days, recommended for CNS a. Peel-away – perfect even block without
tissues and cytological studies (smooth muscles trimming
and skin) b. Plastic ice trays
6. Aniline Oil – clearing embryos, insects and very c. Paper boat – normally utilized for
delicate specimen celloidin embedding
7. Clove oil
8. Carbon tetrachloride – highly toxic (hepatotoxic) F. TRIMMING
9. Methyl benzoate and methyl salicylate  Excess wax is cut off from the block to expose the
tissue surface in preparation for actual cutting
D. IMPREGNATION (INFILTRATION)
 A process whereby the clearing agent is Microtomy
completely removed from the tissue and replaced  Process where embedded tissue is trimmed and
by a medium that will completely fill all the cavities, cut in uniform slices for visualization in the
thereby giving a firm consistency to the specimen microscope
 Melting point of common waxes: 45C, 52C, 56C  Microtome- instrument used
and 58C. 56C- routine work  Uses a cutting tool, steel knife or blade attached
 Lab temp: 20-24C - 54-58C melting point to the microtome
 Lab temp: 15-18C – 50-54C melting point
 Also known as infiltration Three essential part of microtome
1. Block Holder
Three ways by which paraffin wax impregnation and 2. Knife Carrier and Knife
embedding of tissues may be performed: 3. Pawl, Ratchet, Feed Wheel and Adjustment
1. By manual processing Screws
2. By automatic processing
3. By vacuum embedding - It involves wax Kinds of microtome
impregnation under negative atmospheric 1. Rocking microtome (Cambridge)
pressure inside an embedding oven to hasten Invented by Paldwell Trefall in 1881
removal of air bubbles and clearing agent from the Simplest
tissue block. Used to cut small and large paraffin embedded
tissues
Impregnation medium 10-12 u thickness
Paraffin wax Not recommended for serial sections
Celloidin (Colloidon) – bones, teeth, and whole embryo Disadvantages:
2 methods for celloidin method Difficulty of reorienting the block
Wet celloidin – bones, teeth, large brain sections Restrictions in size of tissue block
and organs 2. Rotary microtome (Minot)
Dry celloidin – whole eye section Invented by Minot in 1885-86
Gelatin – histochemical and enzymatic studies To cut paraffin embedded tissues
Plastic Most common type used in routine and research
laboratories
Substitute for Paraffin Wax Can produce ribbons for serial sections
1. Paraplast Cuts in upward vertical motions, flat plane
a. Embeddol Heavier and more stable than rocking microtome
b. Bioloid – embedding eyes Models for cutting ultrathin sections and cryostat
c. Mat use is now available
2. Ester wax Disadvantages:
3. Water soluble waxes (e.g. carbowax) More expensive
Relatively dangerous- knife is in blade
E. EMBEDDING up position
 Also known as casting 3. Sliding microtome
Developed by Adams in 1789
Types of Blocking-out Molds Used Can be used for both celloidin blocks and large
1. Leuckhart’s embedding mold paraffin embedded tissues
2. Compound embedding unit Cuts both small and large tissue with ease

PATRICK R. DE VERA,RMT 10
 HPMTLE REVIEW NOTES

Recommended for cutting hard and rough tissue Specimen is small fixed in Osmium tetroxide,
blocks embedded in plastic
Two types:
Base sledge microtome Care of the Microtome
Two movable pillars holding the 1. Remove all the left over paraffin by using a soft
adjustable knife clamps brush
For cutting celloidin sections 2. After drying the machine and knife holder, wipe
Favored when cutting very hard tissue with xylol
and large blocks 3. Avoid cleaning with xylene
Cutting serial tissues section also 4. Movable portions should be cleaned with oil, to
excellent prevent rusting
Modern models are electrically driven 5. Cover the microtome when not in use
Ideal for resin embedded calcified bone
Standard sliding microtome Microtome knives
Block remains stationary- during 1. Plane-Concave Knife
sectioning Usually 25 mm in length
Ideal for cutting celloidin sections Less concave side is for cutting celloidin
More dangerous because of the embedded tissue blocks
movable knife More concave side is for cutting paraffin
4. Freezing microtome embedded tissue blocks
Invented by Queckett in 1848 2. Biconcave Knife
Stage for block holder is hollow and perforated Usually 120 mm in length
around attached to a flexible pipe where C02 For cutting paraffin embedded sections on a rotary
passes from a cylinder microtome
Knife holder is attached to a lever, connected to a 3. Plane-Wedge Knife
pawl Usually 100 mm in length
Used to cut undehydrated tissues in a frozen state recommended for frozen sections
For rapid diagnosis For hard, tough specimens embedded paraffin
5. Cryostat or cold microtome block in base sledge or sliding microtome
Refrigerated apparatus for fresh tissue microtomy **Knife should be made of good quality steel
**Knife must be able to cut paraffin blocks about 2-4u without any serrations
Freezes the tissue in the block holder for
**Bevel Angle- angle formed between the cutting edge, 27 - 32°. Maintained
facilitation of sectioning by a knife back
Microtome is usually of rotary type **Cutting Angle - 15° (optimum) There is a maximum penetration of tissue
Cold chamber at –50 to 300 C and less distortion
**Clearance angle of 5-100 should be maintained to prevent uneven
Freezes the tissue for 2 to 3 minutes
sections
Cutting sections of 4u
Tissues for fluorescent antibody technique and Honing (Knife Sharpening)
histochemical studies
 Involves the removal of gross nicks on the knife
For intraoperative diagnosis
edge to remove blemishes, and grinding the
cutting edge of the knife on a stone to acquire an
Special Processing technique
even edge
Freeze drying – It is a special way of preserving tissues by
 Removal of gross knick of the knife (coarse
quenching and subsequent dessication of fresh tissues by
honing) to remove blemishes and grinding the
sublimation without the use of any chemical fixative.
cutting edge of the knife on a stone (honing
Time consuming and expensive but has outstanding good
proper) to acquire even edge
features.
Freeze substitution - same with freeze drying but uses the  Degree of sharpness is proportional to the
Rossman’s formula or 1% Acetone and dehydrated with fineness of the abrasive used
absolute alcohol. Alternative: Liquid nitrogen  From Heel to Toe 20 – 30 times
 Direction “ edge first with heel to toe direction”
6. Ultrathin microtome Plane wedge knife- 10- 20 strokes (turned over)
For cutting tissues at 0.5 u Plane concave knife- only the concave surface is
For electron microscopy honed
Uses knife from selected fragments of broken  Mechanical honing- good results, time saving but
plate glass expensive
Types of Hone

PATRICK R. DE VERA,RMT 11
 HPMTLE REVIEW NOTES

1. Belgium yellow-usually gives best result 3. Forceps and squirrel hair brush
2. Arkansas-gives more polishing effect 4. Clean slides 76 x 25 mm, 1 –1.2 mm; frosted ends
3. Fine carborundum - much coarser than above 5. Ice tray
types; used for badly nicked knives 6. Pencil
Precautions during Honing 7. Water bath- set at 100 C below the melting point
Hone should be long enough, (around 8” x 3”)
Hone should be lubricated with warm soapy water or fine oil before use
of paraffin wax
Pressure on the knife should be gentle and steady 8. Drying oven or Hot plate
Hone should be cleaned before, during and after use 9. Forceps and squirrel hair brush
After honing, wipe off the oil or soap from the knife with xylene 10. Clean slides 76 x 25 mm, 1 –1.2 mm; frosted ends
11. Ice tray
Stropping 12. Pencil
 A process whereby the “burr” formed during
honing is removed and the cutting edge of the G. SECTIONING
knife is polished  process whereby tissues are cut into uniformly thin
 Process w/c removes the burr formed during slices or “sections”.
honing
 Purpose is to polish and sharpen the cutting edge 1. Paraffin sections- rocking/rotary
 To polish and sharpen the cutting edge Tissues are embedded in paraffin wax
 From toe to heel Trimmed-excess wax is removed
 Use paddle strop made of horse leather Cut at 4-6u in thickness
 Strops should be oiled on the back (Don’t use Micrometer gauge is set
mineral oil) Knife is tilted at 0-15°complete ribbons are picked
 40 – 120 double strokes up with camel hair brush
 Paddle strop- made of quality horse leather Sections are placed in water bat set at 45-600°C,
 Reverse of honing to flatten the tissues
 Edge last with toe to heel direction Folds and creases maybe removed by stretching
the sections using dissecting needles or forceps
 40-120 strokes double strokes are usually
Ribbons are fished out of the water bath using the
required
Precautions observed in Stropping slides
Knife should be wiped clean with cloth before and after stropping
Pressure during initial stropping should be light 2. Celloidin sections- sliding
Speed stropping should be avoided Sections are placed in water bat set at 45-600°C,
Leather strops are usually dry and require oiling before use (do not use
mineral oil) to flatten the tissues
Vegetable oil, like castor oil should be applied at the back of the strop Folds and creases maybe removed by stretching
Mineral oil is not recommended the sections using dissecting needles or forceps
Stropping surface should be firm and not loose Ribbons are fished out of the water bath using the
Wax should never come in contact with strops
slides
Cut between 10-15u
Disposable Blades
Do not require hardening by chilling before cutting
• sharp cutting edge can cut 2-4u
Sections are cut using sliding microtome
• magnetic knives for cryostat
Cut by WET method, sections and block are kept
moist with 700 alcohol (to avoid dehydration)
Glass Knives
Sections do not come off in ribbons, must be
• trimming and semi-thin sectioning of tissue blocks
collected in alcohol immediately
for electron microscopy
3. Frozen sections- freezing microtome/cryostat
Diamond Knives
Rapid pathologic diagnosis during surgery
• used to cut any type of resin block for electron
Diagnostic and research enzyme histochemistry
microscopy
Diagnostic and research demonstration of soluble
• brittle and inexpensive but durable
substances such as lipids and carbohydrates
Immunofluorescent and immunocytochemical
Other equipments
staining
Some specialized silver stains
1. Water bath- set at 100 C below the melting point
Two methods in Frozen section
of paraffin wax
2. Drying oven or Hot plate
Cold knife procedure

PATRICK R. DE VERA,RMT 12
 HPMTLE REVIEW NOTES

– Knife 40-50 C  Intravital – injected (IV, Intraperitoneal or

– Tissue 5-10 C subcutaneous) (e.g. lithium, carmine or india ink)
– Environment 0-10 C  Supravital – living cells immediately after removal
Cryostat procedure Neutral red – best vital dye
1. Liquid nitrogen Janus green- mitochondria
2. Isopentane cooled by liquid nitrogen
3. Carbon dioxide gas Stains and staining solutions
4. Aerosol spray-quick freezing spray cans of 1. Natural dyes
fluorinated hydrocarbons  Hematoxylin - Hematoxylin campechianum heart
wood, ripening process (oxidizing)
Most common difficulties observed during tissue processing  Cochineal dyes – Coccus cacti -> carmine
(Gregorios, Histopathologic Techniques)  Orcein dyes –lichens (blue or violet colors) (eg.
litmus- indicator)
H. STAINING 2. Synthetic dyes
 The process of applying dyes on the sections to Acidic
see and study the architectural pattern of the Basic
tissue and physical characteristics of the cells Neutral – Romanovsky stains

 A procedure that makes use of heavy metal salts COMMONLY USED STAINING SOLUTIONS USED
which are selectively precipitated on certain Hematoxylin
cellular and tissue components Aluminum hematoxylin
 Ehrlich’s hematoxylin
Affinity (Chemical basis)  Harris’ hematoxylin
 Acidic (Nucleus) – Basic stains  Cole’s hematoxylin
RED: Neutral red, Safranin O, Carmine,  Mayer’s hematoxylin
Hematoxylin Iron hematoxylin
BLUE: MB, Toluidine blue, Celestine blue  Weigert’s
 Basic (Cytoplasm) – Acidic stains  Heidenhain’s
RED: Eosin Y, Eosin B, Phloxine B  Phosphotungstic Acid hematoxylin
YELLOW: Picric acid, Orange G, Rose Bengal Eosin
GREEN: Light green SF, Lissamine green  a red dye
 Routinely used as a counterstain after
Three major groups of staining hematoxylin and before methylene blue
1. Histological staining – dye to tissue
 Stains connective tissue and cytoplasm
2. Histochemical staining –tissue to chem rxn
differentially
3. Immunohistochemical staining - combination
3 forms of Eosin:
1. Eosin Y – yellow (most common)
Methods of staining
2. Eosin B – Erythrosin B
1. Direct
3. Eosin S – alcohol soluble
2. Indirect
 Mordant – link/ “lake” Other stains
 Accentuator – accelerator/ hastens the speed of Acid fuchsin -Picric Acid (Van Gieson’s Stain)- for
staining demonstration of connective tissue
3. Progressive – definite sequence Acridine orange (Masson Stain) - discriminates dead and
4. Regressive –overstained until the desired color is living cells (DNA – green; RNA – red)
obtained Acridine Red 3B - demonstration of calcium salt deposits
5. Metachromatic (Methyl violet or crystal violet, BCB, and phosphatase activities
Safranin, Bismarck brown, Basic fuchsin, MB, Thionine, Alcian blue - water soluble phthalocyanin dye; for connective
Toluidine blue and Azure A, B & C) – tissue components tissue and epithelial mucin
combine w/ these dyes to form a different color from the Aniline blue - cytoplasmic stain; for counterstaining of
surrounding tissue. epithelial section
6. Counterstaining – w/ primary mordant and decolorizer Basic Fuchsin - component of the modified acid fast stain;
7. Metallic Impregnation – not dyes or metallic salts (e.g. plasma stain; for staining of acid fast organisms, for
gold chloride and silver nitrate) mitochondria, for differentiation of smooth muscles
8. Vital stains – living cell constituents Benzidine -for staining hemoglobin

PATRICK R. DE VERA,RMT 13
 HPMTLE REVIEW NOTES

Bismarck brown - used as counterstain for Gram’s Best carmine

technique, for acid fast, for Papanicolau method; used for Langhan’s Iodine
staining diphtheria organism
Carmine - used as chromatin stain for fresh materials in Stains for Protein, Enzymes and Nucleic acids
smear preparations, combined with aluminum chloride to Protein
stain glycogen (Best carmine solution) Alkaline fast green
Celestine blue - used for routine staining of fixed sections, Peracetic acid alcian blue
resistant to strong acids, good nuclear stain
Congo Red - best known as a pH indicator, stains elastic Enzymes
tissues, amyloid, myelin Gomori calcium (ALP)
Crystal violet - a nuclear or chromatin stain, stains amyloid Gomori lead (ACP)
in frozen sections, platelets in blood Lead method for 5-nucleotidase
Giemsa stain - used for staining blood to differentiate WBCs; Metal precipitation of ATPase
blood parasites (pH 7.2 – 7.4)
Gold sublimate - stain used for metallic impregnation, made Nucleic acid
up of gold chloride and mercuric chloride Feulgen technique
Iodine - oldest stain; stain’s amyloid, cellulose, starch, Methyl green pyronin
carotenes, glycogen
Gram’s iodine – stain’s microorganisms and fibrin in tissue Connective tissue stains
sections Reticulin
Lugol’s iodine – used as test for glycogen, amyloid, and PAS
corpora amylacea Gomori’s Silver Impregnation
Janus green B - for demonstrating mitochondria during
intravital staining Collagen
Malachite green - counterstain for Ascaris eggs, Van Gieson’s
erythrocytes and bacterial spore stain, used as a decolorizer Masson’s Trichrome
and counterstain Mallory’s Aniline Blue stain
Methylene blue - stains plasma cells, cytological Azocarmine
examination of sputum for malignant cells, diagnosis of
diphtheria, vital staining of nervous tissue
Neutral red - for demonstration of cell granules and vacuoles Elastic fibers
of phagocytic cells Weigert’s elastic tissue stain
Orcein - stains elastic fibers; Recommended for Orcein (Taenzer Unna Orcein)
dermatological studies; Demonstrates finest and most Krajian’s
delicate fibers in the skin
Osmium tetroxide - used to stain fat (black) Basement membrane
Picric acid - counterstain for acid fuchsin, connective tissues PAS
(Van Gieson’s stain)
Prussian blue -stain for iron Stain for Muscle and Bones
Rhodamine B - fluorescent dye; used with osmic acid to fix Muscle
and stain blood and glandular tissues Modified Gomori’s Trichrome
Silver nitrate - used for identification of spirochetes, reticulin Rapid Gomori Trichrome for Frozen Muscle Tissue
fiber stain Mallory’s Phosphotungstic Acid Hematoxylin
Toluidine blue - used as nuclear stain in fixed tissues; stains Lissamine Fast Red Tartrazine Method (Muscle and Bone)
Nissl granules or chromophilic bodies
Bone
Oil Soluble dyes H and E – Standard
Sudan Black B - most sensitive of the oil soluble dyes; Masson’s Trichrome
greater affinity for phospholipids and neutral fats (TAG) Aniline Blue or Light Green Fiber
Sudan IV (Scharlach R) - most commonly used; stains Schmorl’s Picro thionin
neutral fats but not phospholipids
Sudan III -1st Sudan dye; stains CNS tissue Stains for bone marrow and blood elements
Toluidine blue
Carbohydrate stains Methyl green pyronin
PAS Perl’s
PAS with diastase (Glycogen demonstration) Romanowsky

PATRICK R. DE VERA,RMT 14
 HPMTLE REVIEW NOTES

Wright’s Spirochetes
Giemsa Levaditi’s
Modified steiner and steinier technique
CNS Stains Warthin Starry
Bielschowsky
Bodians Fungal
Sevier Munger Groccot Methenamine Silver
Cresyl fast violet
Weigert Pal Viral stains
Kluver BarreraLuxol Fast Blue Lendrum’s Phloxine Tartrazine Method – viral inclusions
Luxol fast blue PAS Orcein Method (Hepatitis B Surface Antigen)
Weil’s method
Cajal’s gold sublimate Protozoan Stain
Modified PTAH Dilute giemsa - Leishmania, Malaria & Trypanosoma
Modified Holzer
I. MOUNTING
Staining of tissue pigments and deposits • Synthetic water-soluble glycols and resins- used
Endogenous Pigments as mounting media
Perl’s Prussian blue • Preferably tissue blocks are 2-4 mm thick
Turnbull’s Blue reaction Mounting cryostat sections
Benzidine nitroprusside Freezing previously fixed tissue
Modified Fouchet’s Formalin-fixed tissue – cut sections may not adhere to slide
Gmelin’s technique (albumin or chrome glycerin jelly)
Stein’s Iodine Alcohol-fixed tissue – alcohol inhibits freezing (washed 12-
Schmorl’s Ferric Ferricyanide 24 hrs)
Gomori’s aldehyde fuchsin
Mallory’s fuchsin Adhesives
Masson Fontana Technique 1. Mayer’s egg albumin – most common
2. Dried albumin
Calcium deposits 3. Gelatin (1%)
Soluble calcium salts 4. Gelatin- formaldehyde
Gypsum method Oxalate method 5. Starch paste
6. Plasma
Insoluble 7. Poly-L-Lysine
Calcium dye lake reaction 8. APES (3-aminopropylthriethoxysilane)
Metal substitution (Von Kossa’s Silver Nitrate Method)
Mounting medium
Demonstration of copper A. Aqueous medium
Lindquist’s Modified Rhodamine Technique Gelatin, glycerin jelly or gum arabic (solidify)
Glycerol (prevent cracking or drying)
Demonstration of Urates and Pyrophosphates Sugar (increase refractive index) + preservative
Birefringence 1. Water
Lithium carbonate 2. Glycerin (refractive index 1.46)
3. Farrant’s medium (refractive index 1.43)
Staining of microorganisms 4. Apathy’s medium (refractive index 1.52)
Bacteria 5. Brun’s fluid
Gram’s method B. Resinous medium
Gram twort stain 1. Canada Balsam (Refractive index 1.524) – Abus
Brown and Brenn – Nocardia & Actinomyces balsamea
Ziehl Neelsen- Mycobacteria 2. DPX (Refractive index 1.532)
Wade Fite – M.leprae & Nocardia 3. XAM (Refractive index 1.52)
Auramine Rhodamine - Mycobacteria 4. Clarite X (Refractive index 1.544)
Toluidine blue – H.pylori ***Refractive index of glass slide – 1.518
Cresyl violet acetate method – H.pylori
Dieterle method – L.pneumophilia J. LABELLING

PATRICK R. DE VERA,RMT 15
 HPMTLE REVIEW NOTES

DIAGNOSTIC CYTOLOGY - means microscopic GYNECOLOGIC SPECIMEN

examination of cells from different body sites for diagnostic Principle: majority of cervical carcinoma and precancerous
purposes. (e.g, exfoliative cytology and FNA) lesions of the cervix arise from the junction of endocervical/
endocervical mucosa (Transformation zone or T-Zone).
Cytopathology is study of cells in diagnosis of disease. Therefore sampling of T-zone is extremely important for
Exfoliative Cytology – deals with the microscopic study of detection of dysplasia and carcinoma of the cervix.
cells that have been desquamated from epithelial surfaces. Specimen: ideal should include squamous, columnar and
Exfoliated cells may be found in smears that have metaplastic cells.
spontaneously been shed or physically removed from
epithelial and mucous membrane (e.g. vagina, buccal Papanicolau smear (Pap smear) – considered to be the
mucosa and body fluids – sputum, urine, gastric juice and staining method of choice for exfoliative cytology.
CSF). Spontaeous exfoliation is observed in normal cells
due to constant growth and replacement of new cells – more Advantages:
readily observed in malignant tumor cells. Cell samples are 1. Transparent blue staining of cytoplasm is obtained
collected from normally shedding tissues like epithelium. due to the action of high alcoholic content of the
spatula or brush to enhances collection. cytoplasmic counterstain, allowing the overlapping
of cells to be seen and identified.
Recommended for the ff: 2. Excellent nuclear detail to be seen and identified.
1. Detection of malignant cells in body fluids, mainly 3. Color range is predictable and of great value in
used for staging cancer. identification and classification of cells, producing
2. Detection of precancerous cervical lesions in a good differential coloring of basophilic and
women (cervicovaginal smear or Pap’s smeal) acidophilic cells.
3. Assessment of female hormonal status in case of 4. It is valuable in comparing cellular appearance in
sterility and endocrine disorders (Maturation Index smears with their counterpart in similarly stained
(MI)) sections.
4. For the determination of genetic sex (Barr body)
5. For detection of infectious agents Vaginal Hormonal Cytology

Non-Exfoliative: Cells samples collected by needles with Inexpensive and not risky even to pregnant women.
suction pressure. (FNAC) Upper lateral third of the vaginal wall, because it is more
accessible and less likely to be contaminated by cellular
1. Collection and Preparation of Specimen debris or vaginal discharges.
2. Pap smear (cervicovaginal smear)
3. Nipple discharge Cells found in Cervico-vaginal smears:
4. Gastric or Bronchial secretion
5. Pleural and peritoneal fluid 1. Mature superficial cells – polygonal squamous cells
6. Sputum measure 45-50um in diameter. Pale, pink-staining
7. Urine sediment cytoplasm and dark pyknotic nuclei, < 6um.
8. CSF 2. Intermediate cells – medium sized polyhedral; or
elongated cells with basophilic vacuolated cytoplasm.
Smear preparation: name, age, date and type of specime, 3. Parabasal cells – are round to oval cells with small dense
must be made fresh in the doctor’s office basophilic cytoplasm and a total cell diameter of 15-30um.
Fixation: quick immersion to fixative solution or sprayed They are smaller than intermediate cells and have a larger
(~foot distance/ 12 inches away) by the fixative. vesicular nucleus. They are normally found from 2 weeks at
50% alcohol – all types of effusions the age of puberty, after childbirth, with abortions and after
Saccomano fixatives – 50% alcohol and carbowax menopause.
Fluids – more than few drops must be centrifuges (2000RPM
for 2 minutes) or cytospin to the glass slides Other cells that may be found in cervico-vaginal smears:

Common fixatives used for cytologic smears 1. Navicular cells- boat-shaped intermediate cells with the
Equal parts of 95% alcohol and ether – best but has been tendency to fold or curl on edges. Estrogen-progesterone
abandoned X flammability, volatility and fire hazards effect. They are found in the latter half of the menstrual cycle,
95% ethyl alcohol during pregnancy and menopause.
***paper clip 2. Pregnancy cells – round, oval or boat shaped cells with
X striking the bottom of the slide to prevent the dislodging of translucent basophilic cytoplasm observed greatest at the
the cells. center of the cell due to glycogen accumulation, pushing the

PATRICK R. DE VERA,RMT 16
 HPMTLE REVIEW NOTES

nucleus to the side or towards the cell membrane. The Type of specimen: Gastric lavage, gastric brush, and FNA
appearance is characteristic due to a deeper blue stain of (submucosal lesions) – fasting specimen atleast 8 hours
the cytoplasm at the periphery. before washing is performed. Simple irrigation and
3. Endometrial cells – small cells, slightly cylindrical with less aspiration technique
basophilic cytoplasm, occurring in tightly packed groups or Cytologic collection and preparation: same as the respiratory
3 or more. They are found during and 1-10 days after specimens
menstruation will contain to ovarian hormones. A smear
taken during menstruation will contain endometrial cells and Peritoneal, Pleural, and Pericardial fluids
numerous erythrocytes and leukocytes, frequently rendering Jelly like clots forming after removal may be prevented by
the smear unsatisfactory for a reliable assessment. adding 300 units of Heparin for very 100 ml of aspirate.
4. Endocervical cells- occur in large groups or small sheets.
The cytoplasm is usually stained pale blue/gray and is finely Breast Discharge
vacuolated, often with distinct cell borders and nuclei with Principle: Potential detection of malignant cells with clinically
finely granular chromatin. They may present a honeycomb undetected carcinoma (benign breast lesion such as ectasia
appearance when viewed on the end. and Papilloma).
Collection technique: strip the subareolar area and nipple
NON – GYNECOLOGIC SPECIMEN using the thumb and forefinger. Fixed with 95% alcohol or
spray fixative. Site if Left or Right.
Respiratory Tract Specimen
Urinary Tract Specimen
Principle: specimen obtained to exclude the possibility of
malignancy or infectious agents, esp. from patients with Principle: Diagnosis of malignancy, usually of urothelial
immunodeficiency syndrome (e.g. BAL – P. jiroveci origin – prostatic carcinoma (rare)
infection) Type of specimen: voided urine, (M: ok, F: catheterized
specimen is preferred) catheterized specimen, washings
Type of specimen: sputum, BAL, bronchial washings and from bladder of pelvis) 50 ml must be centrifuged
bronchial brushings
Body Cavity effusions
Cytologic collection and preparation
Principle: people with history of cancer or cancer of unknown
Sputum collection (3-4 slides) origin.
Obtained at least consecutive morning sputum specimens Type of specimen: Cavity fluids (pleural, ascetic, peritoneal,
Collect early morning sputum by a deep cough in a wide- pericardial and CSF)
mouthed jar containing Saccomanno fluid (50% ethyl alcohol Cytologic collections and preparations: clean, non-sterile dry
and 2% carbowax). Alveolar macrophage should be container and submitted fresh. Delay in transportation:
streaked with blood or solid particles that should be refrigeration. CSF min of 1cc.
removed.
Sputum vs salivary specimen Fine Needle Aspiration Cytology
Principle: Cancer diagnosis (simple, safe and rapid
Bronchoscopy specimen cytological procedure)
 Bronchial Brushing FNA for palpable masses – breast, thyroid, soft tissue and
 Bronchial Washing lesions (22-23 gauge)
 Bronchial Aspirates FNA for non palpable masses
Cell suspensions (direct taps of pleural or peritoneal
effusions, as well as CSF and synovial fluid). Optimum Slide preparation
amount: 20-30 ml. cells can be viable for up to 4 days if the 1-2 drops using slide-pull technique, max of 4 slides, if there
specimen is kept refrigerated at 4C (do not freeze). is solid tip of the needle is most diagnostic material for
Centrifugation: urine, serous effusions and watery lavages cytological evaluation.
(BAL)
Cytospin slides – 1000 RPM for 1 min (Gregorios) References:
Transbronchial FNA – 1-2 drops, smeared by two –slide pull  Bruce-Gregorios, Jocelyn H., Histopathologic
method and fixed immediately. Techniques 2nd Edition.
 Robbins and Cotran Pathologic Basis of Diseases
Gastrointestinal specimen 8th Edition.
Principle: used to exclude the possibility of malignant tumors

PATRICK R. DE VERA,RMT 17
 HPMTLE REVIEW NOTES

MEDTECH LAWS (3) is a qualified Pathologist, or a duly registered medical

Republic Act No. 5527 technologist of the Philippines with the degree of Bachelor
 June 21, 1969 of Science in Medical Technology/ Bachelor of Science in
 Thirty Two (32) sections Hygiene/ Public Health
 Three (3) amendments R.A. No. 6132, P.D. 498 (4) has been in practice of laboratory medicine or medical
and P.D. 1534. technology for at least ten years prior to his appointment
(5) is not a member of the faculty of
Section 1: Title “Philippine Medical Technology Act of 1969” any medical technology school for at least two (2) years prior
Section 2: Definition of Terms to appointment or having any pecuniary interest direct or
a. Practice of Medical Technology indirect in such institution: Provided, however, That for the
b. Pathologists first three years following the approval of this Act, the
c. Medical Technologists requirements mentioned in number four (4) shall be
d. Medical Laboratory Technicians reduced to five years.
e. Accredited Medical Technology Training
Laboratory Section 9: Executive Officer of the Board
f. Recognized School of Medical Technology Section10: Compensation of the Members of the Board of
g. Council Examiners for Medical Technology
h. Board  P10.00/ each applicant examined
 P5.00/ each applicant granted a certificate of
Section 3: Council of Medical Technology Education, Its registration without examination
Composition
Section 11: Functions and Duties of the Board
Chairman: Director of Higher Education
Vice-Chairman: Chairman of PRC Section 12: Removal of Board Members
Members:  Neglect of duty, incompetency, malpractice or
1. Director of the BRL of the DOH unprofessional, unethical, immoral or
2. Chairman and two (2) members of the Board of dishonorable conduct
Medical Technology
3. Representative of the Deans of Schools of Medical Section 13: Accreditation of School of Medical Technology
Technology and Public Health and Training Laboratories
4. Presidents of the PSP and the PAMET
Section 14: Inhibition Against the Practice of Medical
Section 4: Compensation and Traveling Expenses of Technology
Council a. Duly registered physicians.
 Chairman: P 50.00 b. Medical technologist from other countries called in for
 Members: P 25.00 consultation or as visiting or
exchange professors to colleges or universities: Provided,
Section 5: Functions of the Council of Medical Technology they are only practicing the
Education said function.
c. Medical technologists in the service of the United States
Section 6: Minimum Required Course Armed Forces stationed in the
 4years, 12 months (1 year) internship in Philippines rendering services as such for members of the
accredited laboratories said forces only.

Section 7: Medical Technology Board Section15: Examination

 Chairman: Pathologist  Greater Manila Area, Cebu and Davao
 Members: 2 Registered Medical Technologist  March & August or September
 3 years term
PRC board for Medical Technologists Section 16: Qualification of Examinees
Chairman: Dr. Marilyn A. Cabal –Barza a. Is in good health and is of good moral character;
Members: Ms. Marilyn R. Atienza b. Has completed a course of at least four (4) years leading
Ms. Marian M. Tantingco to the degree of Bachelor of Science in Medical Technology
Section 8: Qualifications of Examiners or Bachelor of Science in Public Health conferred by a
(1) is a Filipino citizen recognized school, college or university in accordance with
(2) is of good moral character this Decree or having graduated from some other profession
and has been actually performing medical technology for the

PATRICK R. DE VERA,RMT 18
 HPMTLE REVIEW NOTES

last five (5) years prior to the date of the examinations, if Section 26: Reinstatement, Reissuance or Replacement of
such performance began prior to June 21, 1969. Certificate

Section 17: Scope of Examination Section 27: Foreign Reciprocity

DAY 1 Section 28: Roster of Medical Technology
Clinical Chemistry 20% Name, address and citizenship of each registered Medical
Microbiology & Parasitology 20% Technologist, date of registration or issuance of certificate,
CM (Urinalysis and other body fluids) 10% and other data which in the opinion of the Board are pertinent
DAY 2
Section 29: Penal Provisions
Hematology 20% Section 30: Separability clause
Blood Banking & Serology 20% Section 31: Repealing Clause
Histopathologic Techniques, Cytotechnology, Medical Section 32: Effectivity
Technology Laws, Related Laws and its implementing
rules, and the Code of Ethics 10% Council of MT education
 No. 3, 4, 5, 6, and 13
Section 18: Report of Ratings
Section 19: Rating in the Examination MT Licensure examination
Section 20: Oath taking  No. 15, 16, 17, 18, 19, 27

Section 21: Issuance of Certificate of Registration MT Regulatory board

 21 years old  No. 7,8, 9, 10, 11, 12, 20, 21, 22, 23, 24, 25, 26,
Laboratory Technician: 29
1. He or she passed the civil service examination for medical
technician given on March 21, SUMMARY OF AMMENDMENTS RA 5527
1964; or
2. Has finished a two-year college course and has at least RA 5527 RA6132 PD 498 PD 1534
SECTION 2 Practice of
one (1) year of experience as medical laboratory technician, Definition of Medical
Provided, that for every year of deficiency in college Terms technology
attainment two (2) years of experience may be substituted; SECTION 3
Council of
Provided, further, that an applicant who has at least ten (10) Medtech Secretary of Commissioner Director of
years of experience as medical laboratory technician as of Chairman Education of PRC CHED
Vice Director of Chairman of the Chairman
the date of approval of this Decree regardless of his Chairman BRL Board of PRC
academic attainment may qualify for registration without SECTION 4
Compensati
examination; or on of Council
3. Has failed to pass the board examination for medical Members P50.00 P50.00
technology but had obtained a general rating of at least 70%. Chairman P50.00**offic P25.00*regardl
Members ial receiving ess of whether
Provided, finally, that a registered medical laboratory salaries from they receive or
technician when employed in the government shall have the the not the regular
government salaries from
equivalent civil service eligibility not lower than second shall not the
grade. receive per government.
diem
SECTION 7
Section 22: Fees Medtech
Board Pathologists Pathologist
Chairman RMT RMT
Section 23: Refusal to issue Certificate of Registration Members President President
 any criminal offense involving moral turpitude Appointed PAMET PRC
3 years for all 3 years for
 any person guilty of immoral or dishonorable Recommend members chairman
conduct ed office 2 years for 1st
member
 unsound mind 1 year for 2nd
 incurable communicable disease member
SECTION 8 Qualificatio
ns of
Section 24: Administrative Investigation – Revocation or Examiners
Suspension of Certificates SECTION 11 Functions and Functions
Duties of the and Duties
Board of Board
Section 25: Appeal

PATRICK R. DE VERA,RMT 19
 HPMTLE REVIEW NOTES

SECTION 13  December 5, 2000

Accreditation DepED DECS DECS
of Schools Medtech board Medtech
Recommend Board QUALIFICATIONS:
ed by: DOH council PRC
of Medtech Medtech board DOH  at least 40 years old
Laboratory Education BRL  With a valid license
Recommend
ed by:  Familiar with the principles and methods of the
SECTITON Actually Actually Actually PRC
16 performing performi performing the
Qualification the practice ng the practice of  At least 5 years executive/ management
for of Medtech practice Medtech for the experience years
Examination for the last 5 of least 5 years
years Medtech
for the Functions of the PRC
last 5
years
SECTION 17 Submission Submission of 1. Administer and implement policies of the
Scope of of schedule schedule of Commission
Examination for subjects subjects is 30
in 4 mos. days
2. Review, revise and approve resolutions
3. Administer and conduct licensure exam of the
*additional various professions
subjects
included: 4. Authorize to require refresher course
Cytotechnology 5. Monitor the performance of schools in licensure
MTL
SECTION 21 examinations
Issuance of Actually Actually Actually 6. Approve and release board exam results
Certificate of performing performi performing the
Registration the practice ng the practice of
7. Issue COR: name, picture, registration number,
of Medtech practice Medtech for the signatures
for the last 5 of least 8 years 8. Recommend to the President the names of
years Medtech
for the *registration professionals for appointment of the various
last 5 without boards
years examination for
medtech = 9. Authorize any officer of the PRC to administer
P115.00 oaths
*registration
without
examination for Professional Regulatory Boards
labtech = • Regulate and monitor the practice of their
P50.00
SECTION 22 respective professions
Fees • Administer investigation
Examination P25.00 P50.00
and – The decision of the PRB shall become
registration P25.00 P25.00 final after 15 days from the receipt of
Registration
notice
without P10.00 P10.00 • Prepare the syllabi of exams
examination • Prepare questions based on these syllabi
Replacemen • Score and rate the exam and submit to the PRC
t Certificate within 10 days from the last day of exam.
SECTION 29 Penal
• Serve as the CPE council
Provisions Republic Act 10912
Approved
date
June
1969
21, August
31, 1970
June 28, 1974 June
1978
11,  “Continuing Professional Development Act of
2016”
OTHER RELATED LAWS  Nineteen (19) sections
“PRC Modernization Act of 2000”  Lapsed into law July 21, 2016
OLD: Presidential Decree 223  Term of Pres. Benigno S. Aquino III
 3 man commission CPD COUNCIL:
 Appointed by the president  Medical Technology Board
 Term office: 9 years CPD providers
NEW: Republic Act No. 8981 REQUIRED CREDIT UNITS: (for all professions)
 3 man commission  60 CU for BS
 Twenty one (21) sections  30 CU for Non-BS
 Appointed by the president REQUIRED CREDIT UNITS (for MT)
 Term office: 7 years  45 CU -> 15 CU

PATRICK R. DE VERA,RMT 20
 HPMTLE REVIEW NOTES

SOURCES of CU License Requirements:

 Seminars  Where?
 Academic preparations  When? (renewal)
 Research  Validity
 Book Authorship  Inspection – every 2 years or as necessary
 Inventions  Changes in the Lab
 Post-graduate trainings  NRL’s
Lab requests and reports:
LICENSURE EXAM REGULATIONS  Consultation between the pathologist and
From PRC Law: physician ( signed) and RMT (signed)
Submission of requirement for application  All reports must be signed by the pathologist
Suspension of examination: before releasing EXCEPT in case of emergency
o Public calamity  Recording: CL reports (1 yr.) and Anatomic &
o Epidemics; 25% of the examinees is forensic records (Permanent)
absent Administrative Order No. 59 s. 2001
Tardiness Section 1: Title: “Rules and Regulations Governing the
o Not > 30 minutes Establishment, Operation and Maintenance of Clinical
o No other examinee has submitted the Laboratories in the Philippines”
paper Section 5: Classification of Laboratories
Classification by Function
 Clinical Pathology
 Anatomic Pathology
Classification by Institutional Character
 Hospital Based Laboratory
 Non-Hospital Based Laboratory
Classification by Service Capability
 Primary: (Routine hematology, Urinalysis,
Fecalysis, for Hospital based: Blood typing &
Quantitative platelet determination ) minimum
working space requirement of 10 m2
 Secondary: (All primary + Routine Chemistry and
Crossmatching) minimum working space
requirement of 20 m2
 Tertiary: (All secondary + Special chemistry,
Special hematology, Immunosero, Microbiology)
minimum working space requirement of 60 m2
Section 7: Requirements and Procedures for Application of
Permit
a. Renewal of License
Region Schedule
NCR Jan to Mar
1,2,3,& CAR Feb to Apr
4,5,& 6 Mar to May
Republic Act 4688 7,8,& 9 Apr to June
 “Philippine Clinical Laboratory Act of 1966” 10, 11,12, CARAGA & ARMM May to July
 Eight (8) sections
 June 18, 1966 National Reference Laboratories
Several implementing rules  NKTI: Hematology & Coagulation
 Scope  RITM: Microbiology& Parasitology
 Classification of lab  Lung Center: Clinical Chemistry
o Function  EAMC: Drug testing
o Service capabilities  SACCL (SLH STD-AIDS Cooperative Center
 Supervision Laboratory): Infectious immunology (Hepatitis B,
 Prescribed number of MT HIV, & HCV)

PATRICK R. DE VERA,RMT 21
 HPMTLE REVIEW NOTES

Republic Act No. 1517 Republic Act No. 9288

 “Blood Banking Act”  “Newborn Screening Act of 2004”
 Six (6) sections  April 7, 2004
 June 16, 1956  Nineteen (19) Sections
Republic Act No. 7719 Test included:
 “National Blood Services Act of 1994” Metabolic disorders
 Fifteen (15) sections  PKU
 May 5, 1994  MSUD
 Voluntarism  Methylmalonic academia
 Classification of Blood Banks  Tyrosinemia
Important provisions  Citrullinemia
 Sections 7, 8, and 9  MCAD deficiency
 Penalties Hormone problems
 Over-pricing—revocation of license to operate  Congenital hypothyroidism
 1-6 mos. In prison  Congenital Adrenal hyperplasia
 5k-50k fine Hemoglobin problems
 No license- 12-20 years, A.O. 9 s. 1995: 6-12  Sickle cell disease
years only  Hemoglobin SC disease
 Transfusing (or failing to discard) contaminated  Beta thalassemia
blood units HIV/ AIDS Prevention and Control Act Others problems
of 1998  Galactosemia
 Cystic Fibrosis
Republic Act No. 8504  Pompe Disease
 “Philippine AIDS Prevention and Control Act of  Spinal Muscle Dystrophy
1998”  Biotidinase deficiency
 February 13, 1998  Severe combined immunodeficiency
 Prevention and control of HIV/AIDS  Mucopolysaccharidosis type 1
 Protecting positive individuals  X –linked adrenoleukodystrophy
 HIV/AIDS info. And Educ. Program Article I: General Provisions
 HIV/AIDS Monitoring system Article II: Definition of Terms
Protection of HIV positive individuals: Article III: Newborn Screening *performed after 24 hrs but not longer
 Considers compulsory HIV testing as unlawful that 3 days
 Any act of discrimination is unlawful Article IV: Implementations
 Social and health services must not be deprived Article V: Final Provisions
 Treating records as confidential
Republic Act No. 11166 Republic Act No. 9165
 “Philippine HIV and AIDS Policy Act”  “Comprehensive Dangerous Act of 2002”
 Fifty seven (57) sections  One hundred two (102) sections
 December 20, 2018  June 7, 2002
 Article I: Philippine National AIDS council  Article I: Definition of terms
o Clandestine lab (illegal manufacture)
 Article II: Information, Education and
o Den/ Dive/ Resort (controlled precursor/
Communication
essential chemicals)
 Article III: Preventive Measures, Safe Practices
o Coddler (protectors)
and Procedures
o PDEA
 Article IV: Screening, Testing and Counseling
 Article II: Unlawful Act and Penalties
Section 29: HIV Testing “15-18 y/o – no need of
 Article III: Dangerous Drug Tests and Record
consent from parent or guardian”
Requirements
 Article V: Health and Support Services
 Article IV: Participation of the Family, Students,
 Article VI: Confidentiality
Teachers and School Authorities in the
 Article VII: Discriminatory Act and Practices and
Enforcement of this Act
Corresponding Penalties
 Article V: Promotion of a National Drug-Free
 Article VIII: Final Provisions
Workplace Program With the Participation of

PATRICK R. DE VERA,RMT 22
 HPMTLE REVIEW NOTES

Private and Labor Sectors and the Department of 2. In the absence of legacy, the physician in-charge
Labor and Employment must look for consent of relatives within 48 hours
 Article VI: Participation of Private and Labor Can a testator cancel the legacy?
Sectors and the Department of Labor and  written statement
Employment  Oral statement with two witnesses
 Article VII: Participation of Local Government  Thru the attending physician
Units
 Article VIII: Program for Treatment and CHED Memorandum Order No. 13 series of 2017
Rehabilitation of Drug Respondents  “Policies, Standards and Guidelines for the
 Article IX: Dangerous Drug Board and Philippine Bachelor of Science in Medical Technology/
Drug Enforcement Agency Medical Laboratory Science (BSMT/ MLS)
 Article X: Appropriation, Management of Funds Program
and Annual Report  Article I: Introduction
 Article XI: Jurisdiction Over Dangerous Drugs  Article II: Authority to Operate
Cases  Article III: General Provisions
 Article XII: Implementing Rules and Regulations  Article IV: Program Specifications
 Article XIII: Final Provisions  Article V: Curriculum
Drug Testing Laboratory  Article VI: Required Resources
Classification  Article VII: Compliance of HEIs
o Screening drug testing lab  Article VIII: Transitory, Repealing and Effectivity
o Confirmatory drug testing lab Provisions
DRUG Analysts/ EAMC Appendix A
Specimen retention:  Article I: Vision & Mission Statement
 5 days for negative specimen and 15 days for  Article II: Description
positive specimen  Article III: Objectives
 Article IV: Requirements
RA 7170
 Article V: General Rules
 ” Organ Donation Act of 1991”
 Article VI: Duties/ Responsibilities of a Clinical
 January 7, 1992 Instructor/ Intern Coordinator/ Clinical Coordinator
 Nineteen (19) sections  Article VII: Duties and Responsibilities of Interns
Definition of terms:
 Article VIII: Merits and Demerits
 Decedent – deceased individual
 Article IX: Responsibilities of the Higher Education
 Donor- individual authorized-> decedent Institution (HEI) and the Accredited Medical
 Testator- individual who makes a legacy of all part Technology Training Laboratory
of his body  Article X: Performance Evaluation
 Recipient  Article XI: Sanctions
 Organ sale
 Legacy CODE OF ETHICS
March 7, 1997 PAMET
What are the organs that can be transplanted? Norma N. Chang
Who can give their parts as a donation?
 Deceased donors Medical Technology Code of Ethics
 Living donors
The donor himself As I enter into the practice of Medical Technology,
The person who decides to donate
At least 18 y/o I shall accept the responsibilities inherent to being a
Any of the following persons (in order of priority) may donate professional;
all or a part of the decedent’s body:
– Spouse I shall uphold the law and shall not engage in illegal work nor
– Son/daughter (18 and above) cooperate with anyone so engaged;
– Either parent
– Siblings (18 and above) I shall avoid associating or being identified with any
– Guardian at the time of death enterprise of questionable character;
Manner of executing a donation:
1. The death must be confirmed first

PATRICK R. DE VERA,RMT 23
 HPMTLE REVIEW NOTES

I shall work and act in a strict spirit of fairness to employer,  ONLY recognized organization for RMTs P.D.
clients, contractors, employees and in a spirit of personal 223, June 22, 1973 creating the PRC.
helpfulness and fraternity toward other members of the  September 15, 1963 – Public Health
profession;
Laboratory, Sta. Cruz, Manila
I shall use only honorable means of competition for PAMET insignia
professional employment or services and shall refrain form  Circle – continuous involvement (practice and
unfairly injuring, directly or indirectly, the professional education -integrated).
reputation, projects or business of a fellow medical  Triangle – love, respect, and integrity
technologist;  Microscope and snake – science of the
medical technology profession
I shall accept employment from more than one employer
 Green – color of health
only when there in no conflict of interest;
 1964 – first PAMET boards was elected
I shall perform professional work in a manner that merits full Core values
confidence and trust carried out with absolute reliability, 1. Integrity
accuracy, fairness and honesty; 2. Professionalism
3. Commitment
I shall review the professional work of other medical 4. Excellence
technologists, when requested, fairly and in confidence 5. Unity
whether they are subordinates or employees, authors of Roster of PAMET Presidents
proposals for grants or contracts, authors of technical papers
or other publications or involved in litigation; 1. Charlamagne T. Tamondong†: 1963-1967
“Emergence of the Profession”
I shall advance the profession by exchanging general 2. Nardito D. Moraleta: 1967-1970,
information and experience with fellow medical technologists “Professional Recognition”
and other professionals and by contributing to the work of 3. Felix E. Asprer†: 1970-1971, 1973-1977,
professional organizations;
“Legislative Agenda”
I shall restrict my praises, criticisms, views and opinions
within constructive limits and shall not use the knowledge I 4. Bernardo T. Tabaosares†: 1971-1973,
know for selfish ends; “Celebration of the Profession”
5. Angelina R. Jose†: 1973, “Career Advocacy”
I shall treat any information I acquired about individuals in
6. Venerable C.V. Oca†: 1977- 1981,
the course of my work as strictly confidential, and may be
divulged only to authorized persons or entities or with “Educational Enhancement”
consent of the individual when necessary; 7. Carmencita P. Acedera: 1982-1991, “Image
Building”
I shall report any infractions of these principles of 8. Marilyn R. Atienza: 1992-1006, “Proactivism”
professional conduct to the authorities responsible of 9. Norma N. Chang: 1997-2000. “International
enforcement of applicable laws or regulations, or to the Leadership”
Ethics Committee of the Philippine Association of Medical 10. Agnes B. Medenilla: 2001-2002, 2005-2006,
Technologists as may be appropriate. To these principles, “Organizational Dynamism”
11. Shirley F. Cruzada: 2003-2004,
I hereby subscribe and pledge to conduct myself at all times
in a manner befitting the dignity of my profession. “Interdisciplinary Networking”
12. Leila M. Florento: 2007-2013, “Global
Philippine Association of Medical Technologists, Perspectives”
Inc. (PAMET) 13. Romeo Joseph J. Ignacio: 2013-2015,
 Crisanto G. Almario – “Father of PAMET” “Golden Celebration”
 1st national convention – FEU, September 20, 14. Ronaldo E. Puno: 2015- present,
1964 “Empowerment”
 1st President - Charlamagne T. Tamondong
 SEC registration – October 14, 1969,
Registration no. 39570

PATRICK R. DE VERA,RMT 24
 HPMTLE REVIEW NOTES

PAMET Hymn Roster of PASMETH Presidents

Music: Francis Jerota Pefanco 1. Dr. Gustavo Reyes: 1970-1973, 1980-1981,
Lyrics: Hector Gentapanan Gayares, Jr. UST
2. Dr. Ibarra T. Panopio: 1973-1974, Velez
Beloved PAMET College
From various lands, races and places 3. Dr. Angelita G. Adeva: 1974-1977, UST
With grateful hearts we blend our voices 4. Dr. Elizabeth M. del Rio: 1977-1980, 1982-
This day our beloved, PAMET 1983, Martinez Memorial Colleges
From whence unity and love cometh, 5. Dr. Claro D. Cabrera: 1981-1982, FEU
We join together in brotherhood 6. Dr. Norma V. Lerma: 1983-1984, UST
To live up to Thine ideals we should 7. Dr. Vicencio T. Torres: 1984-1985, Luzon
In fields of advancement and learning Colleges
Thy noble goals maybe our bearing. 8. Prof. Nardito D. Moraleta: 1985-1988, FEU
Loyal and true we’ll be to thee 9. Dean Norma N. Chang: 1988-1995, San
Beloved PAMET this we say, Juan de Dios Educational Foundation, Inc.
For service to God and humanity 10. Prof. Rodolfo R. Rabor: 1996-2000, UST
With joy we sing for thee till eternity. 11. Dr. Nini F. Lim: 2000-2002, PWU
12. Dr. Zenaida C. Cajucom: 2002-2010, WCC
Philippine Association of Schools of Medical and Martinez Memorial Colleges
Technology and Public Health, Inc. (PASMETH) 13. Dr. Magdalena F. Natividad: 2010-2012, FEU
 May 13, 1970 14. Dean Bernard U. Ebuen: 2012-present,
 SEC registration – October 6, 1985 Arellano University.
 PASMETH is also the founding member of
ASEAN Association of Schools of Medical Medical Technologist’s Prayer
Technology (AASMT)
God, who by calling is to the vocation of a medical
PASMETH logo technologists, has placed upon us the obligation of
Circle – continuity of learning and the never ending being a constant help in the scientific care of the sick,
quest for excellence in the academic field. grant us by Thy divine light a deep insight into the
Diamond - 4 objectives of the association serious responsibility of our tasks.
 To encourage a thorough study of the need By Thy divine wisdom awaken in us growing zeal and
and problems of Medical Technology and determination to increase in our knowledge of how to
Public Health education and to offer solutions search for the unedifying causes of sickness and
to them. disease; how to recognize the evidence of physical
 To work for the continuous development of changes; how to make important chemical analyses,
Medical Technology and Public Health and other valuable tests so helpful in caring for the
education in order that the profession will be sick.
of maximum service of the country. By the divine love permit us in this way to share with
 To take a united stand on matters which affect those who directly care for the sick that thus we may
the interests of medical technology and public be constantly working through the eternal physician
health education. Christ our Lord.
 To seek advice, aid, and assistance from any Amen.
government or private entity for the fulfillment
of the association’s aims and purposes. References:
 CPD units (www.prc.gov.ph)
Microscope – represents the field of Medical  Medical Technology Law and Other related
Technology and Public Health laws (www.lawphil.net)
1970 – year the association was founded  PRC Modernization Act (ched.gov.ph)
 Benitez et al. (2019), Principles of Medical
Laboratory Science 1, C&E Publishing.

PATRICK R. DE VERA,RMT 25