CN109557149A - Digital microcurrent-controlled chip and pathogen immunologic detection method based on pcb board - Google Patents
Digital microcurrent-controlled chip and pathogen immunologic detection method based on pcb board Download PDFInfo
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Abstract
The present invention relates to micro fluidic chip technical fields, more particularly to a kind of digital microcurrent-controlled chip and pathogen immunologic detection method based on pcb board, including upper layer chip, the wall of lower layer chip and connection the two, upper layer chip includes ito glass, lower layer chip includes pcb board basal layer, the workspace that pcb board basal layer is equipped with pin area and is formed by electrode, workspace includes liquid storage tank, drop formation unit, sample cell, analytical unit and waste liquid pool, pcb board basal layer top is equipped with insulating layer, liquid storage tank is connected to by droplet generating unit with analytical unit, sample cell is connected to analytical unit, waste liquid after waste liquid pool is reacted for analytical unit flows into.Above-mentioned digital microcurrent-controlled chip and pathogen immunologic detection method based on pcb board manufactures chip using pcb board, and technical maturity is simply, at low cost, is suitble to commercially produce；Multilayer wiring is, it can be achieved that high density, high-throughput electrode arrangements.
Description
Technical field
The present invention relates to micro fluidic chip technical fields, and in particular to a kind of digital microcurrent-controlled chip based on pcb board and
Pathogen immunologic detection method.
Background technique
Transfusion safety is one of important content of national health care, with the raising of the population of China general level of the health, to defeated
The demand of blood is in continuous upward trend.Many important communicable diseases, such as HBV, HCV, HIV may all borrow blood transfusion way
Diameter is propagated.The blood that stringent screening blood can avoid carrying pathogen is applied to clinical and infects receptor, is to ensure blood transfusion
The effective means of safety.Compared with developed countries, the blood sampling station in China/more dispersed, small scale, equipment is poor, in blood
There is also some places having much room for improvement in detection.In equipment aspect, it is domestic mainly using loading devices such as TECAN EVO and
The imported large-scales detection device such as HAMILTON F.A.M.E24/20 carries out automatic detection.These equipment volumes are huge, and general
The characteristics of center blood station can be mounted on, carry out batch quantity analysis, do not adapt to China's blood sampling point dispersion.
Immune detection is the important technological platform in medical diagnosis on disease.Immunoassay method has very high specific and sensitive
Degree, is widely used in clinical detection.Especially in pathogen detection, although PCR detection method obtain in recent years it is quick
Development, but its defect in terms of specificity does not obtain the improvement of essence, and immune detection is still most reliable, most common inspection
Survey method.But the usual operating process of conventional immunoassay is complicated, needs time-consuming a few hours even one day or more, is difficult to realize fast
Speed detection；A large amount of expensive immunoreagents are expended, and instrument and equipment is complicated, is unsuitable for carrying out field assay.Large biological molecule
Diffusion velocity, the molecular migration time be influence immune response rate key factor.The diffusion coefficient of large biological molecule is general
For 10-5cm2/s.Transit time and diffusion length it is square directly proportional, when migration distance is about 1cm, transit time is that number is small
Up to one day or so；When migration distance is 1um, transit time was less than 1 minute.Relative to routine immunization detection method or lead to
Road declines flow control immune detection.Currently, there is not yet the digital microcurrent-controlled immuno-chip based on pcb board is reported.
Summary of the invention
In order to solve the above technical problems, The present invention provides a kind of digital microcurrent-controlled chip and cause of disease based on pcb board
Body immunologic detection method, technical maturity and simple is at low cost, is suitble to commercially produce；Multilayer wiring is, it can be achieved that high density, height
Flux electrode arrangements can carry out the Multiple detection of multi objective.
In order to reach above-mentioned technical effect, the present invention includes following technical scheme: a kind of based on the digital microcurrent-controlled of pcb board
Chip, the wall including both upper layer chip, lower layer chip and connection, the upper layer chip includes ito glass, the lower layer
Chip includes pcb board basal layer, the workspace that the pcb board basal layer is equipped with pin area and is formed by electrode, the work
Area includes liquid storage tank, drop formation unit, sample cell, analytical unit and waste liquid pool, and pcb board basal layer top is equipped with insulation
Layer, the liquid storage tank are connected to by droplet generating unit with analytical unit, and the sample cell is connected to the analytical unit, described
Waste liquid after waste liquid pool is reacted for analytical unit flows into.
It is perfused with antibody-solutions in the liquid storage tank, sample to be tested is perfused in the sample cell, the waste liquid pool is for anti-
Waste liquid is answered to be discharged into；The analytical unit is reacted and is detected for sample to be tested；The pin area, which is equipped with, connects the digital miniflow
Control the pin of chip and extraneous drop driving circuit, power supply.
Further, the liquid storage tank has 5, the anti-HbsAg antibody-solutions of mouse are perfused respectively, HRP marks anti-mouse secondary antibody,
HRP substrate developing solution, PBS buffer solution, HRP mark anti-human igg secondary antibody；There are two the sample cells, and sample to be tested is perfused respectively.
Preferably, there are two the analytical units, each analysis unit one sample cell of corresponding connection, each analysis unit
Including four detection zones, respectively a, b, c and d detection zone, respectively corresponded on each detection zone electrode be coated with anti-HbsAg antibody,
HIV antigen, HCV antigen and TP antigen.
On the other hand, the present invention provides a kind of pathogen immunologic detection method using above-mentioned digital microcurrent-controlled chip,
The digital microcurrent-controlled chip includes 5 liquid storage tanks, respectively A, B, C, D and E liquid storage tank, and the anti-HbsAg antibody of mouse is perfused respectively
Solution, HRP mark anti-mouse secondary antibody, HRP substrate developing solution, PBS buffer solution, HRP to mark anti-human igg secondary antibody；Include the following steps:
(1) the sample sample drop that sample cell generates enters analytical unit, sufficiently reacts in analytical unit, the drop row after reaction
Enter to waste liquid pool；
(2) D liquid storage tank generates PBS drop, repeats sample sample in step (1) and drips path, washes away non-specific binding serum, more
After secondary repetition, waste liquid is drained into waste liquid pool；
(3) A liquid storage tank generates the anti-HbsAg antibody drop of mouse, enters analytical unit by drop formation unit, soaks repeatedly
PBS drop is generated by D liquid storage tank after profit and carries out rinse；
(4) the secondary antibody drop that B liquid storage tank generates infiltrates analytical unit, the anti-human secondary antibody drop infiltration point that E liquid storage tank generates
Unit is analysed, and is washed with PBS；
(5) C liquid storage tank generates developing solution drop, carries out chromogenic reaction by drop formation unit transportation to analytical unit；
(6) reaction result is taken pictures by CCD, and carries out gray analysis, compared with negative, positive inner quality standard, is made automatically
As a result judge.
The digital microcurrent-controlled chip includes two analytical units, and each analysis unit is corresponding to be connected to a sample cell, often
A analytical unit includes four detection zones, respectively a, b, c and d detection zone, respectively corresponded on each detection zone electrode be coated with it is anti-
HbsAg antibody, HIV antigen, HCV antigen and TP antigen；
A liquid storage tank generates the area d that the anti-HbsAg antibody drop of mouse enters analytical unit in the step (3)；
The area d for the secondary antibody drop infiltration analytical unit that B liquid storage tank generates in the step (4), E liquid storage tank generate anti-human
The area a, b, c of secondary antibody drop infiltration analytical unit；
C liquid storage tank generates developing solution drop in the step (5), is delivered to analytical unit respectively by drop formation unit
A, b, c, d detection zone carry out chromogenic reaction.
By adopting the above technical scheme, including following the utility model has the advantages that provided by the present invention based on the digital microcurrent-controlled of pcb board
Chip and pathogen immunologic detection method manufacture chip using pcb board, and technical maturity is simply, at low cost, is suitble to business metaplasia
It produces；Multilayer wiring is arranged as needed, it can be achieved that high density, high-throughput electrode arrangements；Multiple determination can be carried out, it can be substantially
Degree reduces reagent consumption and can reduce blood station day-to-day operation cost, good market prospect for blood screening.
Detailed description of the invention
Fig. 1 is the structural schematic diagram of the digital microcurrent-controlled chip provided by the present invention based on pcb board；
Fig. 2 is the electrode arrangement schematic diagram of the digital microcurrent-controlled chip provided by the present invention based on pcb board.
In figure,
100, workspace；200, drop formation unit；300, analytical unit；
1, pcb board basal layer；2, insulating layer；3, electrode；4, Teflon layer；5, ito glass；6, wall；7,120P is inserted
Slot；8, sample pool；9, liquid storage tank；10, waste liquid pool；11, detection zone.
Specific embodiment
The present invention is described in further detail below by specific embodiment and in conjunction with attached drawing.
Embodiment:
A kind of digital microcurrent-controlled chip based on pcb board is present embodiments provided, refering to figure, including upper layer chip, lower layer
The wall 6 of chip and connection the two, the upper layer chip includes ito glass 5, and the lower layer chip includes pcb board basal layer
1, the pcb board basal layer 1 is equipped with pin area and the workspace 100 that is formed by electrode, the workspace include liquid storage tank 9,
Drop formation unit 200, sample cell 8, analytical unit 300 and waste liquid pool 10,1 top of pcb board basal layer are equipped with insulating layer
2, production method are as follows: the spin coating 10um thickness SU8-3010 on pcb board basal layer 1 solidifies, forms insulation for whole uv-exposure 15 seconds
Layer.
The liquid storage tank 9 is connected to by droplet generating unit 200 with analytical unit 300, the sample cell 8 and the analysis
Unit 300 is connected to, and the waste liquid after the waste liquid pool 10 is reacted for analytical unit flows into.
It is perfused with antibody-solutions in the liquid storage tank, sample to be tested is perfused in the sample cell, the waste liquid pool is for anti-
Waste liquid is answered to be discharged into；The analytical unit is reacted and is detected for sample to be tested；The pin area, which is equipped with, connects the digital miniflow
Control the pin of chip and extraneous drop driving circuit, power supply.
There are two the walls 6, is divided into the both ends of the digital microcurrent-controlled chip and is located at the two sides of workspace, often
The upper end of a wall is connect with the upper layer chip, and bottom end is connect with lower layer chip.It is being formed by surface of insulating layer again
Spin coating 150um thickness SU8-3050 exposes 80s in SU8 wall area with exposure mask, and ethyl lactate washes away unexposed portion, leaves solid
The SU8 wall of change.
The ito glass bottom surface and the insulating layer top surface are respectively equipped with Teflon layer 4.Sample can be reduced in electrode
On a possibility that sticking, reduce the resistance of liquid drop movement, while advantageously reducing sample room cross contamination.
The pin area is equipped with 120P slot 7, and the pin of pin area passes through 120P slot and the external control of socket connection
So as to the replacement of detection chip, external control signal is convenient to be loaded on each electrode unit circuit.
In the present embodiment, the liquid storage tank has 5, the anti-HbsAg antibody-solutions of mouse are perfused respectively, HRP marks anti-mouse secondary antibody,
HRP substrate developing solution, PBS buffer solution, HRP mark anti-human igg secondary antibody；There are two the sample cells, and sample to be tested is perfused respectively.
There are two the analytical units 300, each analysis unit one sample cell 8 of corresponding connection, each analysis unit packet
Include four detection zones 11, respectively a, b, c and d detection zone, respectively corresponded on each detection zone electrode be coated with anti-HbsAg antibody,
HIV antigen, HCV antigen and TP antigen.
The present embodiment additionally provides a kind of pathogen immunologic detection method using above-mentioned digital microcurrent-controlled chip, including such as
Lower step:
(1) the sample sample drop that sample cell generates enters analytical unit, sufficiently reacts in analytical unit, the drop row after reaction
Enter to waste liquid pool；
(2) D liquid storage tank generates PBS drop, repeats sample sample in step (1) and drips path, washes away non-specific binding serum, more
After secondary repetition, waste liquid is drained into waste liquid pool；
(3) A liquid storage tank generates the anti-HbsAg antibody-solutions of mouse, enters analytical unit by drop formation unit, soaks repeatedly
PBS drop is generated by D liquid storage tank after profit and carries out rinse；
(4) the secondary antibody drop that B liquid storage tank generates infiltrates analytical unit, the anti-human secondary antibody drop infiltration point that E liquid storage tank generates
Unit is analysed, and is washed with PBS；
(5) C liquid storage tank generates developing solution drop, carries out chromogenic reaction by drop formation unit transportation to analytical unit；
(6) reaction result is taken pictures by CCD, and carries out gray analysis, compared with negative, positive inner quality standard, is made automatically
As a result judge.
The digital microcurrent-controlled chip includes two analytical units, and each analysis unit is corresponding to be connected to a sample cell, often
A analytical unit includes four detection zones, respectively a, b, c and d detection zone, respectively corresponded on each detection zone electrode be coated with it is anti-
HbsAg antibody, HIV antigen, HCV antigen and TP antigen；
A liquid storage tank generates the area d that the anti-HbsAg antibody-solutions of mouse enter analytical unit in the step (3)；
The area d for the secondary antibody drop infiltration analytical unit that B liquid storage tank generates in the step (4), E liquid storage tank generate anti-human
The area a, b, c of secondary antibody drop infiltration analytical unit；
C liquid storage tank generates developing solution drop in the step (5), is delivered to analytical unit respectively by drop formation unit
A, b, c, d detection zone carry out chromogenic reaction.
The foregoing is only a preferred embodiment of the present invention, is not intended to restrict the invention, for the skill of this field
For art personnel, the invention may be variously modified and varied.All within the spirits and principles of the present invention, made any to repair
Change, equivalent replacement, improvement etc., should all be included in the protection scope of the present invention.
Claims (10)
1. a kind of digital microcurrent-controlled chip based on pcb board, which is characterized in that including upper layer chip, lower layer chip and connection two
The wall (6) of person, the upper layer chip include ito glass (5), and the lower layer chip includes pcb board basal layer (1), described
The workspace (100) that pcb board basal layer (1) is equipped with pin area and is formed by electrode, the workspace (100) includes liquid storage tank
(9), drop formation unit (200), sample cell (8), analytical unit (300) and waste liquid pool (10), the pcb board basal layer (1)
Top is equipped with insulating layer (2), and the liquid storage tank (9) is connected to by droplet generating unit (200) with analytical unit (300), described
Sample cell (8) is connected to the analytical unit (300), and the waste liquid pool (10) is for the waste liquor stream after analytical unit (300) reaction
Enter.
2. digital microcurrent-controlled chip according to claim 1, which is characterized in that there are two the walls (6), is divided into
The both ends of the digital microcurrent-controlled chip and the two sides for being located at workspace (100), the upper end and the upper layer of each wall (6)
Chip connection, bottom end is connect with lower layer chip.
3. digital microcurrent-controlled chip according to claim 1, which is characterized in that the ito glass bottom surface and it is described absolutely
Edge layer top surface is respectively equipped with Teflon layer (4).
4. digital microcurrent-controlled chip according to claim 1, which is characterized in that the pin area is equipped with 120P slot (7).
5. digital microcurrent-controlled chip according to claim 1, which is characterized in that the liquid storage tank (9) has 5, fills respectively
Infuse the anti-HbsAg antibody-solutions of mouse, HRP marks anti-mouse secondary antibody, HRP substrate developing solution, PBS buffer solution, HRP mark anti-human igg two
It is anti-；There are two the sample cells (8), and sample to be tested is perfused respectively.
6. digital microcurrent-controlled chip according to claim 5, which is characterized in that there are two the analytical units (300), often
A analytical unit one sample cell of corresponding connection, each analysis unit include four detection zones (11), and respectively a, b, c and d is examined
Area is surveyed, is respectively corresponded on each detection zone electrode and is coated with anti-HbsAg antibody, HIV antigen, HCV antigen and TP antigen.
7. a kind of pathogen immunologic detection method using digital microcurrent-controlled chip described in claim 1-6 any one,
It is characterized in that, the digital microcurrent-controlled chip includes 5 liquid storage tanks, respectively A, B, C, D and E liquid storage tank, and it is anti-that mouse is perfused respectively
HbsAg antibody-solutions, HRP mark anti-mouse secondary antibody, HRP substrate developing solution, PBS buffer solution, HRP to mark anti-human igg secondary antibody；Including
Following steps:
(1) sample drop that sample cell generates enters analytical unit, sufficiently reacts in analytical unit, the drop after reaction is drained into
Waste liquid pool；
(2) D liquid storage tank generates PBS drop, repeats sample drop path in step (1), washes away non-specific binding serum, repeatedly weight
After multiple, waste liquid is drained into waste liquid pool；
(3) A liquid storage tank generates the anti-HbsAg antibody drop of mouse, enters analytical unit by drop formation unit, after infiltrating repeatedly
PBS drop is generated by D liquid storage tank and carries out rinse；
(4) the secondary antibody drop that B liquid storage tank generates infiltrates analytical unit, and the anti-human secondary antibody drop infiltration analysis that E liquid storage tank generates is single
Member, and washed with PBS；
(5) C liquid storage tank generates developing solution drop, carries out chromogenic reaction by drop formation unit transportation to analytical unit；
(6) reaction result is taken pictures by CCD, and carries out gray analysis, compared with negative, positive inner quality standard, makes result automatically
Judgement.
8. pathogen immunologic detection method according to claim 7, which is characterized in that the digital microcurrent-controlled chip includes
Two analytical units, each analysis unit one sample cell of corresponding connection, each analysis unit includes four detection zones, respectively
A, b, c and d detection zone, respectively corresponds that be coated with anti-HbsAg antibody, HIV antigen, HCV antigen and TP anti-on each detection zone electrode
It is former；
A liquid storage tank generates the area d that the anti-HbsAg antibody drop of mouse enters analytical unit in the step (3)；
The area d for the secondary antibody drop infiltration analytical unit that B liquid storage tank generates in the step (4), the anti-human secondary antibody that E liquid storage tank generates
The area a, b, c of drop infiltration analytical unit；
In the step (5) C liquid storage tank generate developing solution drop, by drop formation unit be delivered to respectively analytical unit a,
B, c, d detection zone carry out chromogenic reaction.
9. pathogen immunologic detection method according to claim 7, which is characterized in that the digital microcurrent-controlled chip it is exhausted
Edge layer the production method is as follows the spin coating 10um thickness SU8-3010 on pcb board basal layer, whole uv-exposure 15 seconds solidify, shape
At insulating layer.
10. pathogen immunologic detection method according to claim 9, which is characterized in that be formed by surface of insulating layer
Spin coating 150um thickness SU8-3050 again exposes 80s in SU8 wall area with exposure mask, and ethyl lactate washes away unexposed portion, stays
Under cured SU8 wall.
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