阅读说明：本技术 一种功能化生物多层孔隙膜免疫载体、制备方法、应用 (A kind of functionalization biology multilayer hole film immune carrier, preparation method, application ) 是由 李勇 唐玉国 王红梅 段生宝 丁少华 陈晔洲 魏双施 李剑平 于 2019-07-15 设计创作，主要内容包括：本发明提供了一种功能化生物多层孔隙膜免疫载体,包括生物多层孔隙膜、与生物多层孔隙膜结合的特异性抗生物多层孔隙膜抗体、与膜抗体结合的交联抗体。还提供了功能化生物多层孔隙膜免疫载体的制备方法,利用生物多层孔隙膜的免疫原性及多层多孔性,结合抗体-抗原和抗体-抗体特异性结合进行制备。本发明提供的功能化生物多层孔隙膜免疫载体可以用于制备血型相容性检测的免疫载体,进而可以被安装在血型检测卡中进行血型检测。本发明还提供了血型检测卡进行血型检测的方法。本发明提供的用于血型相容性检测的免疫载体可以用于红细胞正定型、反定型、抗体筛检、抗体鉴定等,可实现快速、准确检测红细胞ABO、Rh、MNS、Kell、Kidd等多种血型系统血型。(The present invention provides a kind of functionalization biology multilayer hole film immune carriers, including biological multilayer hole film, the specific antibiont multilayer hole membrane antibody in conjunction with biological multilayer hole film, the cross-linking antibody in conjunction with membrane antibody.The preparation method of functionalization biology multilayer hole film immune carrier is additionally provided, using the immunogenicity and multi-layer porous property of biological multilayer hole film, prepared by binding antibody-antigen and antibody-antibody specific binding.Functionalization biology multilayer hole film immune carrier provided by the invention can be used for preparing the immune carrier of groups compatible detection, and then can be installed in blood type test card and carry out Blood grouping.The present invention also provides the methods that blood type test card carries out Blood grouping.It can be used for red blood cell positive definite form, reverse type, antibody screening, Identification of the antibodies etc. provided by the present invention for the immune carrier of groups compatible detection, it can be achieved that a variety of blood group system blood groups such as quick, accurate detection red blood cell ABO, Rh, MNS, Kell, Kidd.)

1. a kind of functionalization biology multilayer hole film immune carrier characterized by comprising

Biological multilayer hole film；

Specific antibiont multilayer hole membrane antibody, the specificity antibiont multilayer hole membrane antibody and the biological perforated Antigen and antibody specific occurs for gap film in conjunction with the surface to be attached to the biological multilayer hole film；

Cross-linking antibody, the cross-linking antibody are combined together with the specific antibiont multilayer hole membrane antibody coupling；

Wherein, one end Fab of cross-linking antibody can be coupled with the specific antibiont multilayer hole membrane antibody, another end Fab It can be in conjunction with specific antibody.

2. a kind of functionalization biology multilayer hole film immune carrier according to claim 1, which is characterized in that described special Property biology multilayer hole membrane antibody include source of mouse monoclonal antibody, Yang Yuan is mostly anti-, rabbit source monoclonal antibody/more anti-, Ji Yuan are mostly anti-；The cross-linking antibody Including mostly anti-, the anti-sheep of anti-mouse, anti-, anti-chicken is mostly anti-more than mostly anti-, anti-rabbit；It is described to resist with the specificity in conjunction with the end cross-linking antibody Fab Body is source of mouse monoclonal antibody, Yang Yuan is mostly anti-, rabbit source monoclonal antibody/more anti-, Ji Yuan are mostly anti-.

3. a kind of preparation method of functionalization biology multilayer hole film immune carrier, which comprises the following steps:

S1, the grey sheeting cleaned up or wool fabric or real silk fabric or vegetable fibre fabric are cut into round or quadrangular membrane Then diaphragm is immersed in treatment fluid, is impregnated 2-4 hours in 40-70 DEG C of water-bath by piece, biology multilayer hole film is made；

S2, the biological multilayer hole film and specific antibiont multilayer hole membrane antibody are subjected to antigen and antibody specific knot It closes, washing lotion cleaning, then after being coupled with cross-linking antibody, washing lotion cleaning is cleaned again with washing lotion after being closed with confining liquid, is made such as Functionalization biology multilayer hole film immune carrier described in claim 1；

Wherein, the specific antibiont multilayer hole membrane antibody is that antigen-antibody can occur with the biological multilayer hole film The antibody of specific reaction；

The one end the cross-linking antibody Fab can be coupled with the specific antibiont multilayer hole membrane antibody, and another end Fab can be with Specific antibody combines.

4. a kind of biological multilayer hole film immune carrier for groups compatible detection characterized by comprising

Functionalization biology multilayer hole film immune carrier of any of claims 1 or 2；

Erythrocyte blood type antibody or sensitized erythrocyte blood group antigens, by with the functionalization biology multilayer hole film immune carrier On the cross-linking antibody specifically bound and be attached on the functionalization biology multilayer hole film immune carrier；

Wherein, the sensitized erythrocyte blood group antigens are the red blood cell for having specifically bound the blood cell membrane antibody of non-blood group antibody Blood group antigens.

5. the biological multilayer hole film immune carrier according to claim 4 for groups compatible detection, feature exist In, the erythrocyte blood type antibody include IgM type, IgG type, red blood cell ABO, Rh of IgM-IgG type mixed type, MNS, Kell, The blood group antibody of Kidd blood group system, the erythrocyte blood type antibody titer are greater than 1:32.

6. the biological multilayer hole film immune carrier according to claim 4 for groups compatible detection, feature exist It include that sensitization red blood cells of type A antigen, sensitization Type B red cell antigens, sensitization irregularly resist in, sensitized erythrocyte blood group antigens Body screening red cell antigens, sensitized erythrocyte compose cellular antigens.

7. a kind of preparation method of the biological multilayer hole film immune carrier for groups compatible detection, which is characterized in that packet Include following steps:

S1, the grey sheeting cleaned up or wool fabric or real silk fabric or vegetable fibre fabric are cut into round or quadrangular membrane Then diaphragm is immersed in treatment fluid, is impregnated 2-4 hours in 40-70 DEG C of water-bath by piece, biology multilayer hole film is made；

S2, the biological multilayer hole film and specific antibiont multilayer hole membrane antibody are subjected to antigen and antibody specific knot It closes, washing lotion cleaning, then after being coupled with cross-linking antibody, washing lotion cleaning is cleaned again with washing lotion after being closed with confining liquid, is made such as Functionalization biology multilayer hole film immune carrier described in claim 1；

S3, by erythrocyte blood type antibody or sensitized erythrocyte blood group antigens and the functionalization biology multilayer hole film immune carrier On the cross-linking antibody specifically bound, the biology for groups compatible detection as claimed in claim 4 is made Multilayer hole film immune carrier；

Wherein, the specific antibiont multilayer hole membrane antibody is that antigen-antibody can occur with the biological multilayer hole film The antibody of specific reaction；

One end Fab of cross-linking antibody can be coupled with the specific antibiont multilayer hole membrane antibody, and another end Fab can be with Specific antibody combines, and the specific antibody is can be with the erythrocyte blood type antibody or sensitization in conjunction with the end cross-linking antibody Fab Blood group antigen；

The sensitized erythrocyte blood group antigens are the erythrocyte blood type for having specifically bound the blood cell membrane antibody of non-blood group antibody Antigen.

8. the preparation side of the biological multilayer hole film immune carrier according to claim 7 for groups compatible detection Method, which is characterized in that it further include the preparation of step sensitized erythrocyte blood group antigens, it is as follows: with the erythrocyte membrane of non-blood group antibody Antibody is reacted with fresh red blood cell, when 1+ the intensity of agglutination to appear, with hypotonic solution by erythrocyte splitting, is collected by centrifugation and is caused Quick red cell antigens.

9. a kind of blood type test card characterized by comprising

Solid phase plastic carrier, the solid phase plastic carrier include card slot shell and card slot inner casing, the card slot shell and the card Slot inner casing is detachably connected；

Biological multilayer hole film immune carrier described in claim 4-6 any one for groups compatible detection, the use It is located at the bottom of the solid phase plastic carrier card slot shell in the biological multilayer hole film immune carrier of groups compatible detection；

Absorbent wool, positioned at the bottom of solid phase plastic carrier card slot inner casing；

Isolation film is placed on the top of the absorbent wool.

10. blood type test card according to claim 9, which is characterized in that the solid phase plastic carrier prepares material packet Include polyethylene, polyethylene, polystyrene, styrene-butadiene-acrylonitrile；The solid phase plastic carrier, further includes detection hole, The detection hole includes diplopore, three holes, porous；The isolation film includes filter paper, cellulose fibre, fiberglass packing, porous height Fiber mat or test paper made of molecular material；The absorbent wool includes cotton pulp pad, filter paper.

11. blood type test card according to claim 9, which is characterized in that the preservation or detection temperature of the blood type test card Degree is 4-50 DEG C.

12. a kind of detection method of blood group, which comprises the following steps:

Blood group sample to be detected is added in the detection hole of the blood type test card as described in claim 9-11 any one, is put It sets at least 1 minute；Washing lotion is rinsed, and the colour developing situation of detection hole is observed, and it is then positive for developing the color for red, and developing the color for true qualities is then yin Property；

Wherein, the biological multilayer hole film for groups compatible detection in blood group sample to be detected and blood type test card Antigen and antibody specific reaction has occurred in immune carrier, for the positive；Conversely, for feminine gender.

13. functionalization biology multilayer hole film immune carrier of any of claims 1 or 2 is preparing groups compatible testing product In application.

14. functionalization biology multilayer hole film immune carrier of any of claims 1 or 2 is used for detection and analysis cause of disease in preparation Microbial cell antigen-antibody, bacterial cell antigen-antibody, anti-tumor cell antigen antibody product in application.

Technical field

The present invention relates to field of immunodetection, in particular to a kind of functionalization biology multilayer hole film immune carrier, preparation Method and application.

Background technique

Incompatible blood group will lead to serious hemolytic blood transfusion reaction, groups compatible inspection between blood donor and receptor Survey is to ensure that one of most important approach of safe transfusion, compatible (Compatible) i.e. harmonious coexistence refer in patients serum and do not deposit In specific antibody corresponding with the red cell antigens of input, immune response does not occur and is known as compatibility blood transfusion.Otherwise it is known as That is, there is the antibody of specificity corresponding with the red cell antigens of input in incompatible blood transfusion, occur immune anti-in patients serum It answers, the red blood cell inputted in patient body is impaired down to haemolysis.The specific antigen and blood on the surface red blood cell (Red Blood Cell) Antibody in clear determines blood group, and groups compatible detection includes not only the positive reverse type of ABO, RhD blood typing, by blood before transfusing blood Irregular antibody detection, the detection and identification of clinical significant antibody and cross match blood test etc. in person's serum.

The most traditional goldstandard detection method of Blood grouping is test tube method, and test tube method needs cumbersome centrifugation and purge step Suddenly and the preparation of cell suspending liquid.Other conventional blood type testing methods include slide method, micro column gel indirect anti globulin technique (Micrcolumn gel immunoassay, MGIA) and microwell plate solid phase assay etc..Slide detection technique is a kind of honest and clean Valence, quick blood type method.But this method is not sensitive enough to detection poor antigen subgroup, and can only carry out positive definite form inspection It surveys.MGIA can carry out positive definite form and reverse type detection, and can be avoided cleaning step, but fibrinogen in measuring samples Or apparent leukocytosis, then it will lead to false positive results, while needing hatching and centrifugation step, therefore not applicable development is fallen Area afterwards, especially in most developing countries, it has not been convenient to be used as bedside and detect blood group.In addition, raw material gel particle High price monopolization and limited detection flux be also limitation MGIA in the clinical further widely used unfavorable factor with blood station. Microwell plate solid phase assay then improves the sensitivity of detection, but also the same incubation for needing complexity and washing step.Except above-mentioned Outside method, since fiber filter paper has the fiber properties that can be filtered out red blood cell and retain agglutination red blood cell, filter paper is used Extensive report has been obtained in the method for detection blood group.Filter paper detection technique mainly passes through physical absorption and antibody is fixed to filter On paper, limited sorption capacity is not widely used clinically so the sensibility of detection is lower.In addition, IgG class Monoclonal antibody cannot function as sessile antibody and used on this fiber paper, because even there is positive reaction, but be not aggregated, The red blood cell (red blood cell) of positive reaction will be flushed away, and lead to the generation of false negative reaction, while being limited by antibody preservation condition System is not suitable for the use of field contingency condition.

Summary of the invention

For overcome the deficiencies in the prior art, it is an object of the present invention to provide a kind of functionalization biology multilayer hole films Immune carrier, including biological multilayer hole film, by antigen and antibody specific in conjunction with being attached to the biological multilayer hole film Surface specific antibiont multilayer hole membrane antibody, be incorporated in the specific antibiont multilayer hole membrane antibody coupling Cross-linking antibody together, functionalization biology multilayer hole film immune carrier provided by the invention can be anti-by coupling or antibody Original specific binding in conjunction with erythrocyte blood type antibody or sensitized erythrocyte blood group antigens, is examined again to be further used for blood group It surveys；Functionalization biology multilayer hole film immune carrier provided by the invention can also be as needed, and corresponding specificity is selected to catch Antibody is obtained, Immune Selection and combination are immunoreacted or are continued, for detection and analysis pathogenic microorganism cell Antigen-antibody, bacterial cell antigen-antibody, anti-tumor cell antigen antibody and various cell antigen antibodies.

The present invention provides a kind of functionalization biology multilayer hole film immune carrier, comprising:

Biological multilayer hole film；

Specific biological multilayer hole membrane antibody, the specificity antibiont multilayer hole membrane antibody and the biological multilayer Antigen and antibody specific occurs for hole film in conjunction with the surface to be attached to the biological multilayer hole film；

Cross-linking antibody, the cross-linking antibody are combined together with the specific antibiont multilayer hole membrane antibody coupling；

Wherein, the specific antibiont multilayer hole membrane antibody is that antigen can occur with the biological multilayer hole film The antibody of antibody specificity reaction；

One end Fab of cross-linking antibody can be coupled with the specific antibiont multilayer hole membrane antibody, another end Fab energy Enough in conjunction with specific antibody.

Preferably, the specific biological multilayer hole membrane antibody includes that source of mouse monoclonal antibody, Yang Yuan be mostly anti-, rabbit source monoclonal antibody/more Anti-, Ji Yuan is mostly anti-；The cross-linking antibody includes that mostly anti-, the anti-sheep of anti-mouse is mostly anti-, mostly anti-, the anti-chicken of anti-rabbit is mostly anti-；It is described can be with friendship The specific antibody that the end len antibody Fab combines includes source of mouse monoclonal antibody, Yang Yuan is mostly anti-, rabbit source monoclonal antibody/more anti-, Ji Yuan are mostly anti-.

A second object of the present invention is to provide a kind of preparation method of functionalization biology multilayer hole film immune carrier, packets Include following steps:

S1, the grey sheeting cleaned up or wool fabric or real silk fabric or vegetable fibre fabric are cut into round or side Then diaphragm is immersed in treatment fluid, is impregnated 2-4 hours in 40-70 DEG C of water-bath by shape diaphragm, biology multilayer hole film is made；

S2, the biological multilayer hole film and specific antibiont multilayer hole membrane antibody are subjected to antigen and antibody specific In conjunction with the cleaning of, washing lotion, then after being coupled with cross-linking antibody, washing lotion cleaning is cleaned again with washing lotion after being closed with confining liquid, is made Functionalization biology multilayer hole film immune carrier as described above；

Wherein, the specific antibiont multilayer hole membrane antibody is that antigen can occur with the biological multilayer hole film The antibody of antibody specificity reaction；

One end Fab of cross-linking antibody can be coupled with the specific antibiont multilayer hole membrane antibody, another end Fab energy Enough in conjunction with specific antibody.

Preferably, the specific antibiont multilayer hole membrane antibody is final concentration 1-100 μ g/ml with diluted； The cross-linking antibody is final concentration 1-100 μ g/ml with diluted；Wherein, the dilution is 0.01mol/L, PH 7.0- 7.2 phosphate buffer；

Preferably, the treatment fluid is the 0.01mol/L containing 1% bovine serum albumin(BSA), and PH is the phosphate of 7.0-7.2 Buffer；

Preferably, the washing lotion is 0.01mol/L, and PH is the phosphate buffer of 7.0-7.2；

Preferably, it is 5% degreasing of the phosphate buffered saline of 7.0-7.2 that the confining liquid, which is with 0.01mol/L, PH, Milk powder；Wherein, unit % indicates the quality of the solute in 100ml solution.

Third object of the present invention is to provide a kind of immune loads of the biological multilayer hole film for groups compatible detection Body, comprising:

Present invention functionalization biology multilayer hole film immune carrier as described above；

Erythrocyte blood type antibody or sensitized erythrocyte blood group antigens, by immune with the functionalization biology multilayer hole film The cross-linking antibody on carrier is specifically bound and is attached on the functionalization biology multilayer hole film immune carrier；

Wherein, the sensitized erythrocyte blood group antigens be specifically bound non-blood group antibody blood cell membrane antibody it is red Cell blood group antigens.

Preferably, the erythrocyte blood type antibody include IgM type, IgG type, IgM-IgG type mixed type red blood cell A, B, The antibody of Rh, MNS, Kell, Kidd blood group, the erythrocyte blood type antibody titer are greater than 1:32.

Preferably, the sensitized erythrocyte blood group antigens include sensitization red blood cells of type A antigen, sensitization Type B red cell antigens, Sensitization Irregular antibodies screening red cell antigens, sensitized erythrocyte compose cellular antigens.

Fourth object of the present invention is to provide a kind of immune load of the biological multilayer hole film for groups compatible detection The preparation method of body, comprising the following steps:

S1, the grey sheeting cleaned up or wool fabric or real silk fabric or vegetable fibre fabric are cut into round or side Then diaphragm is immersed in treatment fluid, is impregnated 2-4 hours in 40-70 DEG C of water-bath by shape diaphragm, biology multilayer hole film is made；

S2, the biological multilayer hole film and specific antibiont multilayer hole membrane antibody are subjected to antigen and antibody specific In conjunction with the cleaning of, washing lotion, then after being coupled with cross-linking antibody, washing lotion cleaning is cleaned again with washing lotion after being closed with confining liquid, is made Functionalization biology multilayer hole film immune carrier as described above；

S3, erythrocyte blood type antibody or sensitized erythrocyte blood group antigens are immunized with the functionalization biology multilayer hole film The cross-linking antibody on carrier is specifically bound, and the biological multilayer as described above for groups compatible detection is made Hole film immune carrier；

Wherein, the specific antibiont multilayer hole membrane antibody is that antigen can occur with the biological multilayer hole film The antibody of antibody specificity reaction；

One end Fab of cross-linking antibody can be coupled with the specific antibiont multilayer hole membrane antibody, another end Fab energy It is enough with specific antibody in conjunction with, the specific antibody for can in conjunction with the end cross-linking antibody Fab erythrocyte blood type antibody or Sensitized erythrocyte blood group antigens；

The sensitized erythrocyte blood group antigens are the red blood cell for having specifically bound the blood cell membrane antibody of non-blood group antibody Blood group antigens.

Preferably, further include the preparation of step sensitized erythrocyte blood group antigens, specifically prepare sensitized erythrocyte blood group antigens The step of it is as follows: reacted with the blood cell membrane antibody of non-blood group antibody with fresh red blood cell, when 1+ the intensity of agglutination to appear, use is low Erythrocyte splitting is collected by centrifugation and obtains sensitized erythrocyte antigen by osmometer solution.

Preferably, the hypotonic solution is 0.1%-0.7%Nacl solution；

Preferably, the revolving speed of the centrifugation is 12000rpm, centrifugation time 5min.

Fifth object of the present invention is to provide a kind of blood type test cards, comprising:

Solid phase plastic carrier, the solid phase plastic carrier include card slot shell and card slot inner casing, the card slot shell and institute Card slot inner casing is stated to be detachably connected；

Biological multilayer hole film immune carrier as described above for groups compatible detection, it is described to be used for groups compatible Property detection biological multilayer hole film immune carrier be located at the bottom of the solid phase plastic carrier card slot shell；

Absorbent wool, positioned at the bottom of solid phase plastic carrier card slot inner casing；

Isolation film is placed on the top of the absorbent wool.

Preferably, the material for preparing of the solid phase plastic carrier includes polyethylene, polyethylene, polystyrene, styrene-fourth Butadiene-acrylonitrile；The solid phase plastic carrier, further includes detection hole, and the detection hole includes diplopore, three holes, porous；It is described every It include filter paper, cellulose fibre, fiberglass packing, fiber mat or test paper made of porous polymer materials from film；The water suction Cotton includes cotton pulp pad, filter paper.

Preferably, the preservation of the blood type test card or detection required temperature are 4-50 DEG C.

Sixth object of the present invention is to provide a kind of detection methods of blood group, comprising the following steps:

Blood group sample to be detected is added in the detection hole of blood type test card as described above, is placed at least 1 minute；It washes Liquid rinses, and observes the colour developing situation of detection hole, and it is then positive for developing the color for red, and it is then negative for developing the color for true qualities；

Wherein, the biological perforated for groups compatible detection in blood group sample to be detected and blood type test card Antigen and antibody specific reaction has occurred in gap film immune carrier, for the positive；Conversely, for feminine gender.

Preferably, the washing lotion is the buffer containing 0.05%-0.5%Tween-20, and washing lotion pH value is 7.0-7.2, Dosage is 200-1000 μ l.

7th purpose of the invention is to provide a kind of functionalization biology multilayer hole film immune carrier in preparation blood group inspection Survey the application in product.

8th purpose of the invention is to provide a kind of functionalization biology multilayer hole film immune carrier in preparation for examining It surveys and analysis disease includes but is not limited to pathogenic microorganism cell antigen antibody, bacterial cell antigen-antibody, anti-tumor cell antigen antibody Product in application.

In the present invention, the preparation principle of functionalization biology multilayer hole film immune carrier is the specific antibiont multilayer For the Fab section of hole membrane antibody in conjunction with the biological multilayer hole film, Fc sections free outside；One of Fab section of cross-linking antibody Energy binding specificity antibiont multilayer hole membrane antibody is Fc sections free, another Fab section of cross-linking antibody can be in conjunction with more with biology Layer hole film specific antibody forms the biological multilayer hole film immune carrier of functionalization with the specific antibody of kind.

In the present invention, the preparation principle of the biological multilayer hole film immune carrier for groups compatible detection is to first pass through Functionalization biology perforated is prepared in the preparation method of functionalization biology multilayer hole film immune carrier provided by the present invention Gap film immune carrier resists in the cross-linking antibody by combining on functionalization biology multilayer hole film immune carrier with erythrocyte blood type Coupling occurs for body or sensitized erythrocyte blood group antigens or antigen and antibody specific association reaction is made, and can be used for Blood grouping.

Provided by the present invention for groups compatible detection biological multilayer hole film immune carrier, can be realized quickly, The blood group of a variety of blood group systems such as accurate detection red blood cell ABO, Rh, MNS, Kell, Kidd can carry out positive definite form simultaneously, instead determine The detection of the erythrocyte blood types compatibilities such as type, antibody screening, Identification of the antibodies can also need to carry out items selection according to instant detection, With a variety of blood groups and other testing requirements such as be suitble to hospital, family and field urgent.

The present invention utilizes antigen and antibody specific using the immunogenicity and multi-layer porous property of biological multilayer hole film The biology techniques such as affine combination interaction are by grey sheeting, wool fabric, silk cocoon (real silk fabric) or vegetable fibre fabric It is prepared into the hydrophily biology multilayer hole film with biological activity；Hydrophily biology multilayer hole film by with biological multilayer It is successfully prepared energy metaplasia object multilayer hole film immune carrier, function after the coupling of hole film specific antibody in conjunction with cross-linking antibody again Blood group is prepared into after metaplasia object multilayer hole film immune carrier coupling erythrocyte blood type antibody or sensitized erythrocyte blood group antigens The biological multilayer hole film immune carrier of compatibility detection, the biological multilayer hole film immune carrier for groups compatible detection With isolation film, absorbent wool and plastic shell, assembling forms blood type test card in order, detects for groups compatible.It is red thin If born of the same parents react with the antibody on carrier, regardless of whether agglutination, all can by the antibody capture on carrier, when film after washing It is positive findings when taking on a red color.On the contrary, if there is no specific antigen-antibody reaction between antibody on red blood cell and carrier When, it is negative findings that when being washed with washing lotion, red blood cell, which passes through porous biomembrane and siphoned away by absorbent wool, and film shows true qualities, from And it realizes and quickly carries out blood group positive definite form detection.Other groups compatibles are detected, then are by blood plasma to be detected or serum After being added in reacting hole, the sensitized erythrocyte blood group antigens being coupled on the blood group antibody and biological multilayer hole film in blood sample are anti- It answers, reverse type cell or screening cell is added or after washing lotion is rinsed the red blood cell quilt of antigen-antibody reaction occurs for spectrum cell Capture is fixed on punching on porous biomembrane and can't is washed off, and the red of antigen-antibody reaction does not occur for the aobvious red of biological multilayer hole film Cell is flushed through porous film through flushing liquor and is siphoned away by absorbent wool, and biological multilayer hole film film shows true qualities, to realize anti- Sizing detection, antibody screening and Identification of the antibodies.

Compared with prior art, the beneficial effects of the present invention are:

(1) present invention uses biological multilayer hole film from grey sheeting, wool fabric, silk cocoon (real silk fabric), plants Fibres knit equal natural materials, and using the immunogenicity of these natural materials, its corresponding biology multilayer hole membrane antibody is even Connection, then functionalization biology multilayer hole film immune carrier has been prepared into conjunction with cross-linking antibody, it is (enzyme-linked relative to traditional ELISA Immuno absorbence) experiment in 96 orifice plate frostings unordered, non-specific binding, specific biological multilayer hole membrane carrier surface Antigen and antibody specific combines stronger, and site is more clear, and specificity is more preferable.

(2) biological multilayer hole film of the present invention, is multilayer film, therefore red with blood group antigen or sensitization The surface area that cell blood group antibody combines is big, much larger than 96 orifice-plate microporosity surface areas in tradition ELISA experiment, makes reaction sensibility Enhancing.

(3) the biological multilayer hole film immune carrier of groups compatible detection provided by the present invention can be specifically bound Blood group antigens or blood group antibody in blood group sample to be detected, adsorption capacity is high and secured, shortens the time of Blood grouping, improves The sensitivity of Blood grouping.

(4) blood type test card provided by the present invention is to combine solid phase method and agglutination, realizes to groups compatible Detection, the porosity feature of multilayer hole film, which can be realized, directly washes away unreacted red blood cell, while capture as much as possible The single red blood cell that positive reaction occurs, retains the agglutination block of agglutinating reaction, keeps detection more sensitive, special, accurate.

(5) blood type test card provided by the present invention, since native protein has protective effect to biomolecule, so this Main biological active component is all protected in the detection card of invention, and the antigen being coupled on biological multilayer hole membrane carrier is anti- Body, storage temperature can keep stable antigen active and antibody titer at 4-50 DEG C, and the stability for detecting card is strong, save temperature Degree limitation is small, can be stored at room temperature, and is limited by open-air conditions small, is more suitable for using in harsh environment.

(6) in some preferred embodiments of the invention, the isolation film, the water absorption pad, modeling that include in Blood grouping card structure Expect shell and inner casing, matrix is common consumptive material, and easy to maintain, at low cost, availability is high.

(7) in some preferred embodiments of the invention, used isolation film include but is not limited to filter paper, cellulose fibre, Fiber mat made of fiberglass packing or porous polymer materials or test paper, high molecular material performance and porosity can have Effect separating red corpuscle prevents reverse osmosis, makes to detect clear background, accurate quickly to carry out the interpretation of blood group result.

(8) in some preferred embodiments of the invention, used water absorption pad can be cotton pulp pad, filter paper or other fine and close suctions Water-based material can effectively draw surplus liquid, convenient for observing response immediately as a result, shortening the Blood grouping time, 1 minute progress blood POCT (detection immediately) Blood grouping is realized in type judgement.

(9) blood type test card provided by the present invention, used vertical penetration method, can greatly reduce blood chromatography and The time required to cleaning, simplify operating procedure, shortens detection time.

(10) blood type test card provided by the present invention, it is required to be checked when for detecting positive definite form blood group sample to be detected The amount of blood group sample is people's fresh whole blood of 5-50 μ l, and blood volume needed for detecting is few, saves sample size.

(11) in some preferred embodiments of the invention, the inspection on solid phase plastic carrier that provided blood type test card is taken Gaging hole includes but is not limited to diplopore, three holes, porous, can detect red blood cell ABO, Rh, Kell, Kidd etc. as needed and have been found that And blood group system and reverse type, antibody screening and the Identification of the antibodies recognized by international Blood Transfusion Association.

(12) detection method provided by the present invention, result judgement can carry out artificial interpretation using naked eyes, while can also To combine routine spectra detection to be judged automatically.When artificial interpretation, positive findings and negative findings difference are there is redfree, Color difference is obvious, can quick and precisely judge., it can be achieved that the mass of blood group detects when in conjunction with routine spectra detection, to meet Great amount of samples Blood grouping demand.

(13) functionalization biology multilayer hole film immune carrier provided by the present invention may be used also in addition to being used for Blood grouping As needed, the corresponding specificity capture antibody of selection, is immunoreacted, or is continued immune connection and combined, and is used In including but not limited to pathogenic microorganism, bacterium, tumour and various cell antigen antibody immunization experiment detection and analysises.

The above description is only an overview of the technical scheme of the present invention, in order to better understand the technical means of the present invention, And can be implemented in accordance with the contents of the specification, it is described in detail below with some embodiments.A specific embodiment of the invention It is shown in detail by following embodiment.

Detailed description of the invention

The drawings described herein are used to provide a further understanding of the present invention, constitutes part of this application, this hair Bright illustrative embodiments and their description are used to explain the present invention, and are not constituted improper limitations of the present invention.In the accompanying drawings:

Fig. 1 is ABO positive definite form of the present invention and RhD detection principle diagram；

Fig. 2 is ABO reverse type detection principle diagram of the invention；

Fig. 3 is the preparation technology flow chart of functionalization biology multilayer hole film immune carrier of the invention；

Fig. 4 is the preparation process of the multilayer hole film immune carrier for groups compatible detection of the invention

Flow chart；

Fig. 5 is blood type test card structural exploded view of the invention；

Fig. 6 is the ABO positive definite form testing result figure in one embodiment of the invention.

Wherein, in figure: 1, card slot shell；2, card slot inner casing；3, the first detection hole；4, the second detection hole；5, third detects Hole；6, groove.

Specific embodiment

In order to which invention described herein can be more fully understood by, following embodiment is illustrated.It should understand that these embodiments It being merely to illustrate property purpose and is understood not to limit the invention in any way.

First some terms used herein are explained.

" sensitized erythrocyte " refers to incomplete antibody or autoantibody in conjunction with erythrocyte surface antigen or is attached to red thin Make its sensitization on after birth.

" cell erythrocyte " refers to that one group carries most clinical significant blood group antigens, reflects for erythrocyte blood type Fixed more person-portion red blood cells.

" blood cell membrane antibody of non-blood group antibody " refers to blood cell membrane antibody but is not for erythrocyte blood type system Antibody.

In order to which invention described herein can be more fully understood by, following embodiment is illustrated.