CN108445234A - ELISA kit based on IgG4 antibody capture food allergens - Google Patents
ELISA kit based on IgG4 antibody capture food allergens Download PDFInfo
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Abstract
ELISA kit provided by the invention based on IgG4 antibody capture food allergens, including following reagent, are coated with the detection plate of 4 antibody of anti-human igg；Washing buffer；Sample diluting liquid；Standard items or positive quality control product；The food allergen preparation of biotin labeling and 4 preparation of anti-human igg of biotin labeling；The avidin solution of enzyme label；Substrate developing solution；Terminate liquid.The present invention selects IgG4 to detect food allergen, and is the ELISA kit prepared based on prize law, has the characteristics of high sensitivity, favorable reproducibility, and being capable of the large batch of sample of single treatment.Embodiment proves that kit detection sensitivity provided by the invention reaches 1ng/ml.
Description
Technical field
The present invention relates to the technical fields of Allergic skin test kit, and in particular to is based on IgG4 antibody capture food hypersenstivities
Former kit.
Background technology
Anaphylactia incidence accounts for 30% of total world population or more, and the four big of 21 century is classified as by the World Health Organization
One of noninfectious disease.The incidence of anaphylactia significantly rises in recent years, becomes the public health problem of global concern,
Guo Jia et al. is to anaphylactia epidemiological survey the results show that there is 22% people to suffer from anaphylaxis disease in 1,200,000,000 total populations
Disease, such as allergic rhinitis, asthma, conjunctivitis, eczema, food hypersenstivity, drug allergy and serious allergic reaction etc., serious shadow
Ring people’s lives quality and life security.
Food allergen such as egg product, peanut, milk, soya bean, wheat, tree nuts, fish and crust group food, are main
Food allergen.Wherein in crust group food, shrimp allergen is concerned, it was reported that 0.6%~2.8% anaphylactia is suffered from
Person's prawn allergy；People detect at least 13 kinds of IgE binding proteins in shrimp, but tropomyosin is accredited as only master
Anaphylactogen is wanted, relative molecular mass is between 34000~39000；It is reported that tropomyosin is shrimp, crab, oyster, cuttlefish
The important antigen of equal invertebrates, has highly conserved amino acid sequence.In egg allergen, albumen is more easy to draw than yolk
Allergy is played, ovomucoid is main allergen；Its positive rate is up to 35% in children's food hypersenstivity, and allergy of being grown up also is up to
12%.Due to the seriousness and frequently-occurring of peanut allergy reaction, which causes the extensive concern of medical institutions, it was reported that food
In the allergic reaction of object induction, peanut allergy accounts for 10%~47%.Such disease is increasing in recent years, becomes common disease, more
Morbidity is that China's health and economic development field need the significant problem solved.
Currently, patent CN1620504A is disclosed with based on the information design obtained by a part of gene of predetermined substance
Primer carries out PCR, for measuring the method in food whether there is or not predetermined substance, micro constitutent and identification contained in detection food
It is excellent in terms of harmful predetermined substance anaphylactogen, but this method relies on the primer of known detectable substance, but PCR method is multiple
Miscellaneous, detection time is long, of high cost；Patent CN101285840A discloses a kind of detection method of anaphylactogen in food, although should
Method does not disclose relevant detection antibody type in capable of quickly and easily obtaining testing result but the patent, while without public affairs
Open the detection sensitivity and accuracy of this method detection food allergen.In addition, with the molecular weight peaks of flight mass spectrum and collection of illustrative plates shape
Shape carrys out the protein site of identification mark for basic parameter.Anaphylactogen in food is examined by the feature of diagnosis allergy original albumen
Identification is surveyed, but needs special large-scale precision instrument, testing cost is high, and analysis difficulty needs special testing staff and analysis
Personnel.
In conclusion although there are many method for the related food hypersenstivity detection studied at present, it is slow all to there is detection speed,
It is of high cost, the problem of being not suitable for batch detection, and also it is machine-readable to need specific analytical instrument to carry out, and limits food allergen inspection
The development of survey.
Invention content
In view of this, it is an object of the invention to the kit based on IgG4 antibody test food allergens, the kit
With stronger specificity and higher sensitivity.
In order to achieve the above-mentioned object of the invention, the present invention provides following technical scheme：
The present invention provides the ELISA kits based on IgG4 antibody capture food allergens, including following components：
It is coated with the detection plate of 4 antibody of anti-human igg；
Washing buffer；
Sample diluting liquid；
Standard items or positive quality control product；
The food allergen preparation of biotin labeling and 4 preparation of anti-human igg of biotin labeling；
The avidin solution of enzyme label；
Substrate developing solution；
Terminate liquid.
Preferably, the peridium concentration of 4 antibody of the anti-human igg is 0.1~2000 μ g/ml.
Preferably, the washing buffer is PBS buffer solution.
Preferably, the quality of 4 preparation of anti-human igg of the biotin labeling food allergen preparation and biotin labeling is dense
Degree stands alone as 0.01~8000 μ g/ml.
Preferably, 4 preparation of anti-human igg of the biotin labeling is the preparation or biology of the anti-human igg s of biotin labeling
The preparation of the only anti-human igg 4 of element label.
Preferably, the sample diluting liquid is PBS buffer solution.
Preferably, the component of the PBS buffer solution includes the component of following parts by weight：8.0 parts of NaCl, 0.20 part of KCl,
1.16 part Na2HPO4, 0.20 part of KH2PO4With 1000 parts of water；The pH value of the PBS buffer solution is 7.0~7.6.
Preferably, the hydrogen that the sulfuric acid solution or mass concentration that the terminate liquid is 0.5~2.0mol/L are 30%~40%
Sodium hydroxide solution.
Preferably, 1~5000 times of working solution when the avidin solution concentrate of the enzyme label is use.
Preferably, the type of food allergen includes aquatic products category, poultry in the biotin labeling food allergen preparation
Birds, egg-milk, greengrocery, fruits, cereals and nut fruits.
ELISA kit provided by the invention based on IgG4 antibody capture food allergens, including following reagent, coating
There is the detection plate of 4 antibody of anti-human igg；Washing buffer；Sample diluting liquid；Standard items or positive quality control product；Biotin labeling
4 preparation of anti-human igg of food allergen preparation and biotin labeling；The Avidin of enzyme label；Substrate developing solution；Terminate liquid.This
Invention selects IgG4 antibody to detect food allergen for the first time, and is the ELISA kit prepared based on prize law, has spy
It is anisotropic strong, sensitive high feature, and being capable of the large batch of sample of single treatment.Embodiment proves kit inspection provided by the invention
It surveys sensitivity and reaches 2ng/ml.
Further, the ELISA kit provided by the invention based on IgG4 antibody capture food allergens, standard items or
Positive quality control product behaviour IgG4 albumen.The food allergen albumen of biotin labeling can detect in the liquid such as blood plasma, serum
The allergen specificity IgG4 that any food-induced body of oral feeding generates, has the type of detection food allergen more,
The characteristics of having a wide range of application.
Description of the drawings
Fig. 1 is that kit of the present invention is prepared and process for using figure；
Fig. 2 is the standard curve of IgG4 antibody capture food allergens in embodiment 1.
Specific implementation mode
The present invention provides the ELISA kits based on IgG4 antibody capture food allergens, including following reagent：
It is coated with the detection plate of 4 antibody of anti-human igg；
Washing buffer；
Sample diluting liquid；
Standard items or positive quality control product；
The food allergen preparation of biotin labeling and 4 preparation of anti-human igg of biotin labeling；
The avidin solution of enzyme label；
Substrate developing solution；
Terminate liquid.
It includes being coated with the detection plate of 4 antibody of anti-human igg that the present invention, which provides ELISA kit,.4 antibody of the anti-human igg
Peridium concentration be preferably 0.1~2000 μ g/ml, more preferably 0.1~100 μ g/ml.In the present invention, the anti-human igg 4 is anti-
Body is preferably the monoclonal antibody of only anti-human igg 4.In the present invention, the type of the detection plate is preferably transparent polystyrene
Plate and opaque white polystyrene plate.The transparent polystyrene board is for quantitatively detecting；The opaque white
Polystyrene board is used for qualitative detection.
ELISA kit provided by the invention includes washing buffer.In the present invention, the washing buffer is preferably
PBS buffer solution.The PBS buffer solution preferably includes component 8.0 parts of NaCl, the 0.20 part of KCl of following parts by weight, 1.16 parts
Na2HPO4With 0.20 part of KH2PO4With 1000 parts of water；The pH value of the PBS solution is 7.0~7.6.
ELISA kit provided by the invention includes the food allergen preparation of biotin labeling.The biotin labeling
The mass concentration of food allergen preparation is preferably 0.01~8000 μ g/ml, more preferably 0.1~1000 μ g/ml, most preferably
0.1~100 μ g/ml.In the present invention, the type of food allergen includes aquatic products in the biotin labeling food allergen preparation
Category, fowl poultry kind, egg-milk, greengrocery, fruits, cereals or nut fruits.Wherein aquatic products category include fish, shrimp, crab, shellfish,
Clam etc.；Fowl poultry kind includes pork, chicken, beef, mutton；Egg-milk includes egg, milk etc.；Greengrocery include radish, mushroom,
Celery, capsicum, green pepper；Fruits include citrus, orange, banana, apple, Kiwi berry, mango, grape, pineapple, tomato；Cereal
Class includes wheat, oat, rice, corn, soya bean, pea；Nut fruits include pine nut, peanut, fibert, walnut, American pistachios.
In the present invention, the ELISA kit includes sample diluting liquid.The sample diluting liquid is that pH value is 7.0~7.6
PBS buffer solution.The component of the PBS buffer solution includes following parts by weight of component：8.0 parts of NaCl, 0.20 part of KCl, 1.16 parts
Na2HPO4, 0.20 part of KH2PO4With 1000 parts of water.
ELISA kit provided by the invention includes standard items or positive quality control product.The standard items or positive quality control product
For people's IgG4 albumen.
ELISA kit provided by the invention includes terminate liquid.The terminate liquid is the sulfuric acid solution of 0.5~2.0mol/L
Or the sodium hydroxide solution that mass concentration is 30%~40%, the more preferably sulfuric acid solution of 1.0mol/L or mass concentration are
35% sodium hydroxide solution.
ELISA kit provided by the invention includes the avidin solution of enzyme label.The avidin solution of the enzyme label
1~5000 times of working solution when to use.The source of the avidin solution of the enzyme label is not particularly limited, using this field
The avidin solution of enzyme label known to technical staff.The avidin solution of the enzyme label is preferably what HRP was marked
Solution of streptavidin.
ELISA kit provided by the invention includes substrate developing solution.In the present invention, the additive amount of the substrate developing solution
The holes preferably 0.02~0.2ml/, the holes more preferably 0.03~0.16ml/.The substrate developing solution is preferably tetramethyl benzidine
(TMB) substrate solution or 2,2- hydrazines-bis- -3- ethyl benzo thiazole phenanthroline -6- sulfonic acid (ABTS) substrate solution.The tetramethyl connection
The mass concentration of aniline is preferably 0.02~0.05%；The matter of the 2,2- hydrazines-bis- -3- ethyl benzo thiazole phenanthroline -6- sulfonic acid
It is preferably 0.05~0.1mg/ml to measure concentration.
In the present invention, the preparation method of the ELISA kit based on IgG4 antibody capture food allergens, preferably
Preparation including the detection plate for being coated with 4 antibody of anti-human igg.
In the present invention, the preparation method of the detection plate for being coated with 4 antibody of anti-igg preferably includes following steps：
1) 4 monoclonal antibody of anti-human igg is diluted with coating buffer solution, obtains coating buffer；
2) coating buffer that the step 1) obtains is added in the reacting hole of detection plate, 4 DEG C overnight；
3) next day adds confining liquid, is washed again after incubation in step 2) described in once washing after overnight detection plate hole
It washs.
The present invention dilutes 4 monoclonal antibody of anti-human igg with coating buffer solution, obtains coating buffer.In the present invention, the dilution
Method be not particularly limited, using diluted technical solution well-known to those skilled in the art.It is described in the present invention
It is preferably carbonate buffer solution to be coated with buffer solution.In the present invention, the concentration of the carbonate buffer solution is preferably 0.01~
0.2mol/L.The pH value of the carbonate buffer solution is preferably 8.5~9.5.
In the present invention, the concentration of the coating buffer is preferably 0.1~2000 μ g/ml, more preferably 0.1~100 μ g/ml.
After obtaining coating buffer, the coating buffer is added in the reacting hole of detection plate by the present invention, and 4 DEG C overnight.The present invention
In, the volume of coating buffer is preferably 0.02~0.2ml coating buffers in each reacting hole.
In the present invention, described 4 DEG C preferably carry out under conditions of standing overnight, and it is firm to be conducive to 4 monoclonal antibody of anti-human igg
Ground is coated in detection plate.
Next day after overnight detection plate hole described in once washing, adds confining liquid, is washed again after incubation.The present invention couple
There is no limit using the technical solution of washing well-known to those skilled in the art for the method for the once washing.This hair
In bright, the additive amount of the washing buffer is preferably at least the volume of coating buffer；The number of the once washing is preferably 2~
6 times；The incubation time of the single wash is preferably every time 0~5min (similarly hereinafter).
After the once washing, confining liquid is added in the detection plate after once washing, is washed again after incubation.The present invention
In, the confining liquid is preferably that mass concentration is the PBS buffer solution containing 1%~5% balf serum albumin.The confining liquid
Additive amount be preferably at least the volume of coating buffer.In the present invention, the time of the incubation is preferably 10~200min.It is described to incubate
The temperature educated is preferably 10~40 DEG C, more preferably 15~35 DEG C.
After the incubation, the present invention is washed detection plate again, in the present invention, the method washed again with
The method of once washing described in above-mentioned technical proposal is identical, and details are not described herein.
It is described wash again after, after the present invention preferably drains the moisture in detection plate naturally, with vacuum bag sealing detection plate,
Obtain the detection plate for being coated with 4 antibody of anti-human igg.
The detection method of ELISA kit based on IgG4 antibody capture food allergens, includes the following steps：
1) sample to be tested, standard items, negative controls are respectively added in the detection plate for being coated with 4 antibody of anti-human igg,
It carries out carrying out first time washing after being once incubated；
2) the food allergen preparation or biology of biotin labeling are added in the detection plate into the step 1) after washing
4 antibody preparation of anti-human igg of element label, carries out washing for second after secondary incubation；
3) Avidin that enzyme marks is added in the detection plate into the step 2) after washing or the strepto- that enzyme marks is affine
Plain solution carries out third time washing after being incubated three times；
4) to substrate developing solution will be added in the detection plate after washing in the step 3), after four times are incubated, to detection plate
Middle addition terminate liquid, the detection plate to be developed the color；
5) the color developing detection plate obtained according to the step 4), obtains qualitative or quantitative result.
Sample to be tested is added in the detection plate for being coated with 4 antibody of anti-human igg by the present invention, and first is carried out after primary incubation
Secondary washing.In the present invention, the sample to be tested includes the liquid containing IgG4 antibody such as serum, blood plasma.The serum or blood plasma
The dilution of equal liquid is 1~32000 times.Dilute serum, blood plasma are preferably dilution in the kit with solution.
In the present invention, in order to keep testing result accurately credible, blank control wells, negative control hole, standard are preferably also set up
Sample wells or positive quality control hole, and ensure that the amount of liquid of each reacting hole addition is consistent.Sample is preferably added in the blank control wells
Product dilution；The liquid samples such as serum or the blood plasma of no allergies crowd are preferably added in the negative control hole；The standard
4 albumen of human IgG is preferably added in sample wells or positive quality control hole.
In the present invention, the temperature being once incubated is preferably 10~40 DEG C, more preferably 15~35 DEG C.It is described once to incubate
The time educated is preferably 10~200min.
In the present invention, the first time wash temperature is preferably 10~40 DEG C, more preferably 15~35 DEG C；The washing is slow
The additive amount of fliud flushing is preferably at least the volume of coating buffer；The number of the once washing is preferably 2~6 times；The single is washed
The incubation time washed is preferably every time 0~5min (similarly hereinafter).
The food allergen preparation of biotin labeling is added in detection plate after being washed to the first time, after secondary incubation
Second is carried out to wash.In the present invention, the additive amount of the food allergen preparation of the biotin labeling is preferably 0.02~
The holes 0.2ml/, the holes more preferably 0.03~0.16ml/.
In the present invention, the temperature of the secondary incubation is preferably 10~40 DEG C, more preferably 15~35 DEG C.It is described secondary to incubate
The time educated is preferably 10~200min.
In the present invention, the method for second of washing is not particularly limited, and use is well-known to those skilled in the art
Washing methods.
After described second is washed, the avidin solution of enzyme label is added into obtained detection plate by the present invention, incubates three times
Third time washing is carried out after educating.In the present invention, the avidin solution of the enzyme label is diluted before use, described diluted
Preferably 1~5000 times of multiple.In the present invention, the additive amount of the avidin solution of the enzyme label is preferably 0.02~0.2ml/
Hole, the holes more preferably 0.03~0.16ml/.
In the present invention, the temperature being incubated three times is preferably 10~40 DEG C, more preferably 15~35 DEG C.It is described to incubate three times
The time educated is preferably 10~200min.
In the present invention, the method for the third time washing is not particularly limited, and use is well-known to those skilled in the art
Washing methods.
After washing three times, substrate developing solution is added into obtained detection plate by the present invention, after four times are incubated, into detection plate
Add terminate liquid, the detection plate to be developed the color.
In the present invention, the additive amount of the substrate developing solution is preferably the holes 0.02~0.2ml/, more preferably 0.03~
The holes 0.16ml/.
In the present invention, the temperature of four incubations is preferably 10~40 DEG C, more preferably 15~35 DEG C.The incubation
Time is preferably 1~45min, more preferably 10~30min.
In the present invention, the additive amount of the terminate liquid is preferably the holes 0.02~0.2ml/, more preferably 0.03~0.16ml/
Hole.
The detection plate of the colour developing obtained described in observation or detection, obtains qualitative or quantitative result.
In the present invention, the qualitative results are colors in the reacting hole for observe by the naked eye detection plate, in blank control wells
It does not develop the color with negative control hole, in the case that positive quality control hole is developed the color, sample well colour developing is so judged as that measuring samples are sun
Property；Conversely, in the case where blank well and negative control hole do not develop the color the colour developing of positive quality control hole, sample well does not develop the color, then
Measuring samples are feminine gender.
In the present invention, the quantitative result is judged by detecting the OD values of solution in detection plate reacting hole, with blank
The OD values in each hole are surveyed after control wells zeroing, and quantitative detection is carried out based on standard curve.It is described detection preferably when with
ABTS develops the color, then Detection wavelength is set as 405nm；When TMB develops the color, then Detection wavelength is set as 450nm.
In the present invention, 4 preparation of anti-human igg and standard items of the biotin labeling and the inspection for being coated with 4 antibody of anti-human igg
Drafting board prepares standard curve using double antibody sandwich method.The standard curve is four parameter Logistic Fitting curve equations:Y=
(A-D)/[1+(X/C)B]+D；Wherein A=1.22771；B=-0.82579；C=3.87090；D=0.15802；R2=
0.98616.The ELISA kit detection range of IgG4 antibody capture food allergens is 0.488~2000ng/ml.
The kit provided by the invention based on IgG4 antibody capture food allergens is carried out with reference to embodiment detailed
Thin explanation, but they cannot be interpreted as limiting the scope of the present invention.
Embodiment 1
Coating：It is 2.0 μ g/ that specific anti-human IgG4 monoclonal antibodies, which are diluted to protein content, with carbonate buffer solution
The coating buffer of ml.0.05ml coating buffers are separately added into each reacting hole of polystyrene board, 4 DEG C overnight.Next day discards in hole
0.35ml washing buffers are added per hole for solution, wash 3 times, are incubated 1.5min (referred to as washing, similarly hereinafter) every time；
Closing：The PBS buffer solution that 0.05ml contains 2% balf serum albumin, pH are separately added into each reacting hole：7.2
(confining liquid), 15~35 DEG C of incubation 120min, discards solution in hole, washing is same as above；
Be added gradient dilution 4 albumen of human IgG (standard sample wells, concentration be shown in Table 1), it is PBS buffer solution (blank control wells), non-
Allergy volunteers sera (negative control hole) and sample to be tested, that is, suspicious egg, milk and Wheat Dood Allergy patients serum (sample well,
Serum dilutes 100 times with PBS buffer solution, is shown in Table 2).15~35 DEG C of incubation 60min, are washed out；
Add detection antibody：The anti-human igg s Dan Ke of 0.05ml biotin labelings are added per hole for standard sample wells and blank control wells
Grand antibody (a concentration of 2000ng/ml), negative control hole and sample well are separately added into the egg of corresponding biotin labeling per hole
Clear anaphylactogen crude extract, the detecting milk allergen crude extract of biotin labeling and the Wheat Dood Allergy original crude extract of biotin labeling, 15
~35 DEG C of incubation 60min, are washed out；
The avidin solution of enzyme label：The Avidin that 0.05ml horseradish peroxidase-labeleds are separately added into per hole is molten
Liquid, 15~35 DEG C of incubation 30min, is washed out；
Substrate solution is added to develop the color：The tmb substrate solution 0.05ml of Fresh is separately added into per hole, 15~35 DEG C are protected from light incubation
15min；
Terminate reaction：It is separately added into 2mol/L sulfuric acid solutions 0.05ml per hole；
Detect OD values：ELISA Plate is placed in microplate reader, the absorbance value (OD at Detection wavelength 450nm450) (table 1), and
Draw standard curve.The each Kong Jun of this experiment does multiple holes, and experiment is repeated 3 times.
The standard concentration and its OD of the ELISA kit of 1 human IgG of table, 4 antibody capture food allergen450Value
| Serial number | Standard items (ng/ml) | OD450(mean value) |
| 1 | 2000 | 1.2587 |
| 2 | 1000 | 1.2260 |
| 3 | 500 | 1.2058 |
| 4 | 250 | 1.1890 |
| 5 | 125 | 1.1680 |
| 6 | 62.5 | 0.9866 |
| 7 | 31.25 | 0.9780 |
| 8 | 15.63 | 0.9020 |
| 9 | 7.813 | 0.9290 |
| 10 | 3.906 | 0.7316 |
| 11 | 1.953 | 0.5201 |
| 12 | 0.977 | 0.4424 |
| 13 | 0.488 | 0.3031 |
| 14 | 0 | 0.1734 |
Standard curve is drawn according to the concentration of the IgG4 standard items of serial number 1,3,5,7,9,11,13 and 14 and OD values, is such as schemed
2, four parameter Logistic Fitting curve equations are obtained according to standard curve:Y=(A-D)/[1+ (X/C)B]+D；Wherein A=
1.22771；B=-0.82579；C=3.87090；D=0.15802；R2=0.98616.
It can be obtained by embodiment 1：(1) the ELISA kit detection range of IgG4 antibody captures food allergen is
0.488~2000ng/ml；(2) sensitivity of the ELISA kit of IgG4 antibody captures food allergen is 1ng/ml.
The case where table 2 human IgG, 4 antibody capture food anaphylactogen specificity IgG 4 antibody
As seen from the above embodiment, the ELISA kit provided by the invention based on IgG4 antibody capture food allergens,
IgG4 antibody test food allergens, and the ELISA kit prepared based on prize law are selected, has detection sensitivity strong,
The characteristics of accuracy is high, favorable reproducibility, and being capable of the large batch of sample of single treatment.Embodiment proves reagent provided by the invention
Box detection sensitivity reaches 1ng/ml.Type with detection food allergen is more, the characteristics of having a wide range of application.
The above is only a preferred embodiment of the present invention, it is noted that for the ordinary skill people of the art
For member, various improvements and modifications may be made without departing from the principle of the present invention, these improvements and modifications are also answered
It is considered as protection scope of the present invention.
Claims (10)
1. the ELISA kit based on IgG4 antibody capture food allergens, which is characterized in that including following components：
It is coated with the detection plate of 4 antibody of anti-human igg；
Washing buffer；
Sample diluting liquid；
Standard items or positive quality control product；
The food allergen preparation of biotin labeling and 4 preparation of anti-human igg of biotin labeling；
The avidin solution of enzyme label；
Substrate developing solution；
Terminate liquid.
2. kit according to claim 1, which is characterized in that the peridium concentration of 4 antibody of the anti-human igg be 0.1~
2000μg/ml。
3. kit according to claim 1 or 2, which is characterized in that the washing buffer is PBS buffer solution.
4. kit according to claim 1, which is characterized in that the biotin labeling food allergen preparation and biology
The mass concentration of 4 preparation of anti-human igg of element label stands alone as 0.01~8000 μ g/ml.
5. kit according to claim 1, which is characterized in that 4 preparation of anti-human igg of the biotin labeling is biology
The preparation of the anti-human igg s of element label or the only anti-human igg 4 of biotin labeling.
6. kit according to claim 1, which is characterized in that the sample diluting liquid is PBS buffer solution.
7. the kit according to claim 3 or 5, which is characterized in that the PBS buffer solution includes the group of following parts by weight
Point：8.0 parts of NaCl, 0.20 part of KCl, 1.16 parts of Na2HPO4, 0.20 part of KH2PO4With 1000 parts of water；The pH of the PBS buffer solution
Value is 7.0~7.6.
8. kit according to claim 1, which is characterized in that the terminate liquid is that the sulfuric acid of 0.5~2.0mol/L is molten
The sodium hydroxide solution that liquid or mass concentration are 30%~40%.
9. kit according to claim 1, which is characterized in that a concentration of use of the avidin solution of the enzyme label
When 1~5000 times of working solution.
10. kit according to claim 1, which is characterized in that eaten in the biotin labeling food allergen preparation
The type of object anaphylactogen includes aquatic products category, fowl poultry kind, egg-milk, greengrocery, fruits, cereals or nut fruits.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201810413738.9A CN108445234A (en) | 2018-05-03 | 2018-05-03 | ELISA kit based on IgG4 antibody capture food allergens |
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201810413738.9A CN108445234A (en) | 2018-05-03 | 2018-05-03 | ELISA kit based on IgG4 antibody capture food allergens |
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| CN108445234A true CN108445234A (en) | 2018-08-24 |
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|---|---|---|---|
| CN201810413738.9A Pending CN108445234A (en) | 2018-05-03 | 2018-05-03 | ELISA kit based on IgG4 antibody capture food allergens |
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Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101256186A (en) * | 2008-03-28 | 2008-09-03 | 杭州浙大生科生物技术有限公司 | Food anaphylactogen specificity IgG4 antibody ELISA detection kit and preparation method thereof |
| WO2009082798A1 (en) * | 2008-01-02 | 2009-07-09 | Fundação De Amparo À Pesquisa Do Estado De Minas Gerais - Fapemig | Method and kit for quantifying of allergen-especific human igg subclasses for the control and attendance of specific immunotherapy |
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Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2009082798A1 (en) * | 2008-01-02 | 2009-07-09 | Fundação De Amparo À Pesquisa Do Estado De Minas Gerais - Fapemig | Method and kit for quantifying of allergen-especific human igg subclasses for the control and attendance of specific immunotherapy |
| CN101256186A (en) * | 2008-03-28 | 2008-09-03 | 杭州浙大生科生物技术有限公司 | Food anaphylactogen specificity IgG4 antibody ELISA detection kit and preparation method thereof |
Non-Patent Citations (2)
| Title |
|---|
| E.E.GUHSL等人: "IgE,IgG4 and IgA specific to Bet v 1-related food allergens do not predict oral alergy syndrome", 《ALLERGY》 * |
| V. OLIVIERI等: "Capture assay for specific IgE An improved quantitative method", 《JOURNAL OFLMMUNOLOGICAL METHODS》 * |
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