CN109001466A - It is a kind of for the pulmonary surfactant protein kit of OSA and its application - Google Patents

Abstract

The present invention provides a kind of pulmonary surfactant protein kits for OSA, pulmonary surfactant protein for OSA includes from the SP-A and SP-D detected in serum and the SP-B and SP-C that detect from serum or lung-douching fluid, there are four compartments for kit, first compartment stores the detection serum of SP-A and SP-D to be detected, second compartment stores the serum or lung-douching fluid of SP-B and SP-C to be detected, third compartment accommodates the reagent for detecting Curosurf, and the 4th compartment accommodates the reagent for promoting the reagent in third subregion to detect.Kit detection time of the present invention shortened in one day, it can even accomplish that morning acquisition obtains testing result blood afternoon, substantially reduce detection time, accelerate treatment process, and it without an example mistaken diagnosis or is failed to pinpoint a disease in diagnosis in 86 patients of test, diagnostic accuracy has reached 100%, has wide applicability in OSA diagnostic field.

Description

A kind of pulmonary surfactant protein test kit and application thereof for OSA

technical field

The present invention relates to medical technical field, specifically, the present invention is specifically related to a kind of technical field of medical kit.

Background technique

Both obstructive sleep apnea (OSA) and hypertension (HTN) are thought to increase the risk of cardiovascular events and all-cause mortality. In middle-aged male patients, the clinical overlap is very common and further increases the risk of cardiovascular disease time. With the deepening of research in recent years, the range of target organ damage in patients with OSA complicated with HTN has been continuously expanded, and new possible mechanisms of action have been continuously proposed. However, the diagnostic methods need to be further improved in order to achieve more accurate and rapid diagnosis and treatment.

Since Veergaard first proposed pulmonary surfactant in 1929, we have known the exact composition of pulmonary surfactant and the synthesis, secretion, regulation and metabolism of pulmonary surfactant. The main function of pulmonary surfactant is to reduce alveolar surface tension and prevent end-expiratory lung collapse, which plays an important role in maintaining the normal structure of alveoli and stabilizing ventilation. The research on pulmonary surfactant has been mainly focused on acute respiratory distress syndrome (ARDS) and neonatal respiratory distress syndrome for a long time. With further research, it is found that it not only exists in the alveoli, but also exists in the trachea and all levels. In the bronchi, surfactants in the respiratory tract can maintain airway stability, promote fluid removal, prevent fluid accumulation in the lumen, and have important functions such as host defense and immune regulation, but the application of pulmonary surfactant to obstructive sleep There are few reports in the diagnosis of apnea (OSA), and the existing diagnosis results are not satisfactory, the repeatability is poor, and the diagnostic accuracy and diagnostic speed are very poor.

Contents of the invention

However, there are few reports on the application of pulmonary surfactant to the diagnosis of obstructive sleep apnea (OSA) in the prior art, and the existing diagnosis results are not satisfactory, the repeatability is poor, and the diagnostic accuracy and diagnostic speed are very poor. technical problem, the present invention provides a lung surfactant protein kit for OSA, using blood samples as diagnostic materials, using the kit of the present invention to detect SP-A, SP-B, SP-C and SP-D It can quickly carry out diagnosis and treatment, and has wide applicability in the field of OSA treatment.

The present invention provides a pulmonary surfactant protein kit for OSA, wherein the pulmonary surfactant protein for OSA includes SP-A, SP-B, SP-C and SP-D.

In the present invention, SP-A and SP-D are detected from serum, and SP-B and SP-C are extracted and detected from lung lavage fluid.

In the present invention, SP-A, SP-D, SP-B and SP-C are detected from serum.

In the present invention, the kit has four compartments, the first compartment stores the detection serum of SP-A and SP-D to be detected, and the second compartment stores the serum of SP-B and SP-C to be detected or lung lavage liquid, the third compartment contains reagents for detecting pulmonary surfactant, and the fourth compartment contains reagents for facilitating detection of the reagents in the third compartment.

In the present invention, the reagents used to detect pulmonary surfactant are ethylenediaminetetraacetic acid, enzyme-labeled reagents and SP-A, SP-D, SP-B and SP-C standard substance dilutions.

In the present invention, the reagents used to facilitate the detection of the reagents in the third partition are distilled water and the substrate TMB.

In the present invention, the detection method in the kit includes the following steps:

(1) Blood samples are anticoagulated with 3ml of ethylenediaminetetraacetic acid, and the anticoagulated blood samples or lung lavage fluid are centrifuged to remove sediment particles to obtain serum or clarified lung lavage fluid, then subpackage the samples and seal them in- Store in refrigerator at 80°C;

(2) Before the test, take out the thawed serum or clarified lung lavage fluid from the refrigerator at room temperature for 1 hour, put it in a centrifuge after thawing and centrifuge at 300 rpm for 20 minutes, collect the supernatant and coat it with the purified corresponding index antibody Microwell plate, made of solid-phase antibody;

(3) Add the corresponding indicators SP-A, SP-B, SP-C and SP-D to the microwells coated with monoclonal antibodies in sequence, and then combine with the corresponding indicator antibodies labeled with HRP to form antibody-antigen-enzyme-labeled antibody complexes ;

(4) After thorough washing, the substrate TMB is added for color development;

(5) Measure the absorbance OD value with a microplate reader at a wavelength of 450nm, and calculate the SP-A, SP-B, SP-C and SP-D concentrations in the sample by a standard curve.

In the present invention, the kit includes a box body and an enzyme-labeled plate, and two sets of enzyme-labeled plates are arranged at the bottom of one side of the box body.

In the present invention, the box body is provided with a first compartment, a second compartment, a third compartment and a fourth compartment, and a group of enzyme-labeled plates are respectively provided at the bottom of the first compartment and the second compartment, and the enzyme-labeled There is a sealing plate on the coating plate, centrifuge tube holes, pipette holes and sealing plate height adjustment rods are arranged on the upper part of the first compartment and the second compartment, centrifuge tubes are arranged in the centrifuge tube holes, pipette holes A pipette is arranged in the middle, reagent bottle holes and pipette holes are arranged in the upper part of the third compartment and the fourth compartment, a washing bottle is arranged in the fourth compartment, and an ear washing ball is arranged in the upper part of the third compartment.

In the present invention, handles are provided on both sides of the box body adjacent to the enzyme-labeled coated plate, four hooks are provided at the end of the box body away from the enzyme-labeled coated plate, a pull ring is provided at the end of the sealing plate, and the reagent bottle hole and Labels are provided next to the centrifuge tube holes.

Application of a diagnostic kit for diagnosis of SP-A, SP-B, SP-C and SP-D concentrations in serum or lung lavage fluid in an OSA diagnosis and treatment device.

Beneficial effects of the present invention:

A lung surfactant protein kit for OSA provided by the invention, the lung surfactant protein used for OSA includes SP-A, SP-B, SP-C and SP-D species, and the detection time of this application is shortened to one day It is even possible to collect blood in the morning and get the test results in the afternoon, which greatly shortens the test time and speeds up the treatment process, and none of the 86 patients in the test was misdiagnosed or missed, and the diagnostic accuracy reached 100%, which is unique in the field of OSA diagnosis Broad applicability.

Description of drawings

Fig. 1 is a schematic diagram of the structure of the kit of the present invention.

In Figure 1, 1-cassette, 2-enzyme label coated plate, 3-sealing plate, 4-pull ring, 5-positioning edge, 6-first compartment, 7-second compartment, 8-third Compartment, 9-the fourth compartment, 10-handle, 11-pipettor, 12-sealing plate height adjustment rod, 13-reagent bottle hole, 14-label, 15-wash bottle, 16-ear wash ball, 17 -Centrifuge tube.

Detailed ways

Below in conjunction with accompanying drawing 1 and embodiment the specific embodiment of the present invention is described in further detail, but method of the present invention is not limited to following embodiment.

Enzyme-coated plate, sealing plate, pipette, label, washing bottle, centrifuge tube for ear washing ball, microplate reader, SP-A, SP-B, SP-C and SP-D standards used in the present invention solution, ethylenediaminetetraacetic acid, enzyme-labeled reagents, distilled water, and substrate TMB can all be purchased from the market.

Embodiment one: the present invention is used for the pulmonary surfactant protein kit of OSA

The present invention provides a pulmonary surfactant protein kit for OSA, wherein the pulmonary surfactant protein for OSA includes SP-A, SP-B, SP-C and SP-D.

In the present invention, SP-A and SP-D are detected from serum, and SP-B and SP-C are extracted and detected from lung lavage fluid.

In the present invention, the kit has four compartments, the first compartment stores the detection serum of SP-A and SP-D to be detected, and the second compartment stores the serum of SP-B and SP-C to be detected or lung lavage liquid, the third compartment contains reagents for detecting pulmonary surfactant, and the fourth compartment contains reagents for facilitating detection of the reagents in the third compartment.

In the present invention, the reagents used to detect pulmonary surfactant are ethylenediaminetetraacetic acid, enzyme-labeled reagents and SP-A, SP-D, SP-B and SP-C standard substance dilutions.

In the present invention, the reagents used to facilitate the detection of the reagents in the third partition are distilled water and the substrate TMB.

Embodiment two: the present invention is used for the pulmonary surfactant protein kit of OSA

The invention provides a lung surfactant protein kit for OSA, said lung surfactant protein for OSA includes SP-A, SP-B, SP-C and SP-D, SP-A, SP- D, SP-B and SP-C detected from serum.

In the present invention, the kit has four compartments, the first compartment stores the detection serum of SP-A and SP-D to be detected, and the second compartment stores the serum of SP-B and SP-C to be detected or lung lavage liquid, the third compartment contains reagents for the detection of pulmonary surfactant, the fourth compartment contains reagents for facilitating the detection of reagents in the third compartment, the reagent for detection of pulmonary surfactant is ethylenediamine Tetraacetic acid, enzyme-labeled reagents and SP-A, SP-D, SP-B and SP-C standard dilutions, reagents used to facilitate the detection of reagents in the third partition are distilled water, substrate TMB.

Embodiment three: the present invention is used for the pulmonary surfactant protein kit of OSA

In the present invention, the detection method in the kit includes the following steps:

(1) Blood samples are anticoagulated with 3ml of ethylenediaminetetraacetic acid, and the anticoagulated blood samples or lung lavage fluid are centrifuged to remove sediment particles to obtain serum or clarified lung lavage fluid, then subpackage the samples and seal them in- Store in refrigerator at 80°C;

(2) Before the test, take out the thawed serum or clarified lung lavage fluid from the refrigerator at room temperature for 1 hour, put it in a centrifuge after thawing and centrifuge at 300 rpm for 20 minutes, collect the supernatant and coat it with the purified corresponding index antibody Microwell plate, made of solid-phase antibody;

(3) Add the corresponding indicators SP-A, SP-B, SP-C and SP-D to the microwells coated with monoclonal antibodies in sequence, and then combine with the corresponding indicator antibodies labeled with HRP to form antibody-antigen-enzyme-labeled antibody complexes ;

(4) After thorough washing, the substrate TMB is added for color development;

(5) Measure the absorbance OD value with a microplate reader at a wavelength of 450nm, and calculate the SP-A, SP-B, SP-C and SP-D concentrations in the sample by a standard curve.

Embodiment four: the present invention is used for the pulmonary surfactant protein kit of OSA

In the present invention, the detection method in the kit includes the following steps:

(1) Specimen storage: First, anticoagulate the blood specimen with 3ml of ethylenediaminetetraacetic acid (EDTA), centrifuge it to remove a large number of precipitated particles in the blood specimen or lung lavage fluid, then divide the samples and seal them in a -80°C refrigerator For storage, avoid repeated removal of samples prior to concentration determination.

(2) Pre-measurement treatment: Take out the freezer box from the laboratory refrigerator and place it indoors for 1 hour to fully thaw, put it in a centrifuge and centrifuge at 300 rpm for 20 minutes, and collect serum or lung lavage fluid supernatant .

(3) Solid-phase antibody preparation: Take out the enzyme-labeled coated plate, set 10 standard wells on this microplate, repeatedly add or take out the standard and standard diluent into the ten wells, and put the final standard well The sample volume is 50μl, and the concentrations in the ten wells are 18ng/L, 18ng/L, 12ng/L, 12ng/L, 6ng/L, 6ng/L, 3ng/L, 3ng/L, 1.5ng/L, 1.5ng/L.

(4) Adding samples: Set the sample wells to be tested and the blank control wells in the solid-phase antibody-coated microwells, add 40 μl of sample diluent and 10 μl of the sample to be tested into the above-mentioned sample wells to be tested, and the operation process Note that the sample should be added to the bottom of the well, and try to keep the pipette tip from touching the bottom of the well. No treatment was done in the blank control wells. Shake the enzyme-labeled plate to mix the liquid as much as possible, seal it with plate sealing film, and place it in an incubator for 30 minutes at a temperature of 37°C. At this time, the sample forms an antigen-antibody complex with the corresponding index antibody.

(5) Enzyme addition: Remove the sealing plate, take out the enzyme-labeled coated plate, discard the liquid in the well and dry it, add the concentrated washing solution diluted 30 times with distilled water to each well, and let it rest for 30 seconds Discard the liquid again, repeat the process of adding washing liquid, standing still, and discarding five times, then add 50 μl of enzyme-labeled reagent to each well except the blank control well, put it back into the incubator, take it out and add washing liquid, the operation steps are the same as above. At this time, an antigen-antibody-enzyme-labeled reagent complex is formed.

(6) Color development and termination reaction: Add 50 μl of substrate TMB to each well of the enzyme-labeled plate for color development, mix well and let it stand in the dark for 15 minutes. At this time, the liquid in the well turns blue, add 50 μl of stop solution, The liquid turns from blue to yellow after acidification, and the depth of yellow is proportional to the concentration of the sample in the well.

(7) Determination of the calculated concentration: within 15 minutes of adding the stop solution, the microplate reader measures the absorbance value of the liquid in each well, i.e. the OD value, zeros the blank control well, and the wavelength of the microplate reader is controlled at 450nm. Combined with the standard concentration and absorbance value, the actual concentration of the sample is calculated through the standard curve and regression equation.

Embodiment five: the present invention is used for the pulmonary surfactant protein kit of OSA

In the present invention, the kit includes a box body 1 and an enzyme-labeled coated plate 2, and two sets of enzyme-labeled coated plates 2 are arranged at the bottom of one side of the box body 1.

In the present invention, the box body 1 is provided with a first compartment 6, a second compartment 7, a third compartment 8 and a fourth compartment 9, and the bottoms of the first compartment 6 and the second compartment 7 are respectively provided with a group of enzymes A standard coating plate 2, a sealing plate 3 is arranged on the enzyme label coating plate 2, a centrifuge tube hole, a pipette hole and a sealing plate height adjustment rod 12 are arranged on the upper part of the first compartment 6 and the second compartment 7, A centrifuge tube 17 is arranged in the centrifuge tube hole, a pipette 11 is arranged in the pipette hole, a reagent bottle hole 13 and a pipette hole are arranged on the top of the third compartment 8 and the fourth compartment 9, and the fourth compartment 9 is provided with a washing bottle 15, the upper part of the third compartment 8 is provided with an ear washing ball 16, the box body 1 is provided with handles 10 on both sides adjacent to the enzyme label coating plate 2, and the box body 1 is far away from the enzyme label coating plate 2 There are four hooks at one end, a pull ring 4 is provided at the end of the sealing plate 3, and a label 14 is provided next to the reagent bottle hole 13 and the centrifuge tube hole.

Application of a diagnostic kit for diagnosis of SP-A, SP-B, SP-C and SP-D concentrations in serum or lung lavage fluid in an OSA diagnosis and treatment device.

Embodiment 6: The effect verification test of the pulmonary surfactant protein kit for OSA of the present invention

A total of 86 male patients who were hospitalized in the hypertension diagnosis and treatment center from December 2014 to December 2017 and underwent sleep apnea monitoring in the sleep monitoring center were selected. The selected patients had a clear history of snoring (self, family members or patients in the same ward) Nocturnal snoring), unexplained daytime sleepiness, unexplained lip and tongue cyanosis, micrognathia, and obesity whose blood pressure is difficult to control. Index = weight (Kg)/height (m) ² , and lung function, lung volume and pulse shock detection were performed.

Diagnostic criteria: Obstructive sleep apnea refers to the disappearance of oronasal airflow, while thoracic and abdominal breathing still exists. Apnea occurs due to upper airway obstruction, but the respiratory drive function of the central nervous system is normal, and continues to issue breathing exercise commands to excite the respiratory muscles, so the thoracoabdominal breathing exercise still exists.

Sleep apnea: During sleep, the airflow of oronasal breathing disappears or is significantly weakened (compared with the baseline amplitude by >90%), and the duration is ≥10S. Hypopnea: Oral and nasal airflow decreased by >30% compared with the baseline level during sleep, accompanied by a decrease in SaO ₂ ≥ 4%, and lasted for ≥ 10 seconds; or a decrease in oral and nasal airflow by > 50% compared with the baseline level, accompanied by a decrease in SaO ₂ ≥ 3% , duration ≥ 10S. Abnormal breathing efforts with duration ≥ 10 seconds and associated micro-awakenings were also recorded as hypopnea events if the criteria for apnea or hypopnea were not met. According to the apnea hypopnea index, it is divided into mild (5 times/min≤AHI<15 times/min), moderate (15 times/min≤AHI<30 times/min), severe (AHI≥30 times/min) OSAS .

All selected patients were measured for bust, abdominal circumference, weight, height, etc. Bust measurement refers to the length displayed by a soft tape measure around the fullest part of the breast, and abdominal measurement refers to the length displayed by a soft tape measure around the navel. 1. Ask the patient to take off his shoes and socks and wear unlined clothes when measuring height. Blood pressure measurement standard: the operator first checks whether the instrument is in good condition and is in a standby state, and checks the patient's name. Before taking blood pressure measurement, you should empty your bladder and rest for at least 20 minutes, and try to sit in unlined clothing. First ask the patient to keep the correct posture quietly, fully expose the entire right arm, turn on the switch of the mercury column sphygmomanometer, tie the cuff to two horizontal fingers above the patient's elbow joint, the tightness should be able to fit 1 finger between the cuff and the skin, At this time, the operator wears the stethoscope, palpates the most obvious part of the arterial pulse in the cubital fossa and covers the stethoscope body, closes the valve of the inflatable ball of the stethoscope and starts to inflate, looks directly at the scale of the stethoscope, and continues to inflate after the arterial pulse sound disappears during auscultation make the mercury column rise. Close the valve of the inflatable ball before squeezing the ball and then inflate until the arterial pulsation disappears, continue to add about 25-30mm Hg, open the stethoscope air valve to deflate slowly and evenly, and pay attention to the arterial pulsation sound, when the arterial pulsation first appears The systolic pressure is the systolic pressure, and the diastolic pressure is when the last pulse of the artery or is significantly weakened. After the measurement, the cuff is removed to exhaust the air, and the sphygmomanometer is tilted at a certain angle to turn off the mercury column switch. Repeat the above method to measure blood pressure 3 times, and take the average value.

The determination of biochemical indicators was completed by the hospital testing center. All the included subjects fasted for 12 hours. In the morning of the next day, the patient took a sitting position to draw 3-5ml of blood from the median venous vein of the elbow, and treated it with 3ml of ethylenediaminetetraacetic acid (EDTA) for anticoagulant treatment. Determination of special and routine biochemical indicators. Routine biochemical indicators (cholesterol, triglycerides, fasting blood glucose, electrolytes, glycosylated hemoglobin, high-sensitivity C-reactive protein, etc.) were measured by Hitachi 7600 automatic biochemical analyzer.

Utilize the pulmonary surfactant protein kit for OSA of the embodiment of the present invention five and the method for embodiment four to measure SP-A in collecting serum, SP-B, SP-C and SP-D concentration.

parameter non-OSA group OSA group p age) 45.22±6.17 45.23±5.55 0.781 BMI (kg/m ² ) 28.17±3.02 28.09±2.89 0.049 SP-A(ng/ml) 158.17±32.22 141.45±32.44 0.242 SP-B(ng/ml) 47.81±5.14 39.56±5.42 <0.001 SP-C(ng/ml) 35.01±4.76 34.19±5.21 0.572 SP-D (ng/ml) 19.24±3.76 17.69±3.47 0.147

From the above data, it can be seen that judging from one or both of SP-A, SP-B, SP-C and SP-D is affected by the test error and the patient's own function, and it is impossible to accurately determine whether there is OSA. , but if there is a test box that can perform four tests at the same time, it can accurately and quickly determine whether the patient has OSA. From the perspective of 86 test subjects in the parallel test, the test time of this application is shortened to within one day, and it can even be achieved The blood was collected in the morning and the test results were obtained in the afternoon, which greatly shortened the test time and accelerated the treatment process. Moreover, none of the above-mentioned 86 cases were misdiagnosed or missed, and the diagnostic accuracy reached 100%.

As mentioned above, the present invention can be better realized. The above-mentioned embodiment is only a description of the preferred implementation of the present invention, and does not limit the scope of the present invention. Various modifications and improvements made by technicians to the technical solution of the present invention shall fall within the scope of protection determined by the present invention.

Claims (10)

1. a kind of pulmonary surfactant protein kit for OSA, which is characterized in that the lung surface active egg for OSA White includes SP-A, SP-B, SP-C and SP-D.

2. being used for the pulmonary surfactant protein kit of OSA as described in claim 1, which is characterized in that the SP-A and SP- D extracts detection from detection, SP-B and SP-C in serum from lung-douching fluid.

3. being used for the pulmonary surfactant protein kit of OSA as described in claim 1, which is characterized in that described SP-A, SP- D, SP-B and SP-C are detected from serum.

4. being used for the pulmonary surfactant protein kit of OSA as described in claim 1, which is characterized in that the kit There are four compartment, first compartment stores the detection serum of SP-A and SP-D to be detected, and second compartment stores SP-B and SP- to be detected The serum or lung-douching fluid of C, third compartment accommodate the reagent for detecting Curosurf, and the 4th compartment, which accommodates, to be used In the reagent for promoting the reagent detection in third subregion.

5. being used for the pulmonary surfactant protein kit of OSA as claimed in claim 4, which is characterized in that described for detecting The reagent of Curosurf is ethylenediamine tetra-acetic acid, enzyme marking reagent and SP-A, SP-D, SP-B and SP-C standard dilutions.

6. being used for the pulmonary surfactant protein kit of OSA as claimed in claim 4, which is characterized in that described for promoting The reagent of reagent detection in third subregion is distilled water, substrate TMB.

7. the pulmonary surfactant protein kit for OSA as described in claim 5 or 6, which is characterized in that the reagent Detection method includes the following steps: in box

(1) blood specimen is anticoagulant with ethylenediamine tetra-acetic acid 3ml, the blood specimen or lung-douching fluid centrifugal treating by anticoagulation with Removal precipitating particle obtains serum or clarification lung-douching fluid, then dispenses sample, seals up for safekeeping into -80 DEG C of refrigerator storages；

(2) it is taken out before test from refrigerator and places 1h defrosting serum or clarification lung-douching fluid at room temperature, be put into centrifuge after defrosting It is centrifuged 20 minutes with 300 revs/min of revolving speed, the corresponding index antibody for collecting supernatant purifying is coated with microwell plate, and solid phase is made Antibody；

(3) phase of corresponding index SP-A, SP-B, SP-C and SP-D again with HRP label is sequentially added into the micropore of coating monoclonal antibody It answers indicator antibody to combine, forms antibody-antigene-hrp-antibody complex；

(4) add substrate TMB colour developing after thoroughly washing；

(5) absorbance OD value is measured under 450nm wavelength with microplate reader, SP-A, SP-B in sample are calculated by standard curve, SP-C and SP-D concentration.

8. being used for the pulmonary surfactant protein kit of OSA as claimed in claim 7, which is characterized in that the kit packet Box body and enzyme mark coating plate are included, one side bottom of box body is equipped with two groups of enzyme marks and is coated with plate.

9. being used for the pulmonary surfactant protein kit of OSA as claimed in claim 8, which is characterized in that the box body is equipped with First compartment, second compartment, third compartment and the 4th compartment, first compartment and second compartment bottom are respectively equipped with one group of enzyme mark packet By plate, enzyme mark, which is coated with above plate, is equipped with sealing plate, first compartment and second compartment top be equipped with centrifugation pore, pipette well and Sealing plate height regulating rod is centrifuged in pore and is equipped with centrifuge tube, and pipette, third compartment and the 4th compartment are equipped in pipette well Top is equipped with reagent bottle hole and pipette well, and the 4th compartment is equipped with wash bottle, and third upper compartment is equipped with ear washing bulb, box body and enzyme mark It is coated with the adjacent two sides of plate and is equipped with handle, box body is set far from one end of enzyme mark coating plate there are four hook, and sealing plate end is equipped with Pull ring.

10. SP-A in serum or lung-douching fluid is utilized as described in claim 1, SP-B, the diagnosis of SP-C and SP-D concentration Application of the kit in OSA diagnosing and treating apparatus.