CN1811454A - Cyclomycin family antibiotic enzyme-linked immunoassay reagent kit - Google Patents
Cyclomycin family antibiotic enzyme-linked immunoassay reagent kit Download PDFInfo
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- CN1811454A CN1811454A CN 200610037948 CN200610037948A CN1811454A CN 1811454 A CN1811454 A CN 1811454A CN 200610037948 CN200610037948 CN 200610037948 CN 200610037948 A CN200610037948 A CN 200610037948A CN 1811454 A CN1811454 A CN 1811454A
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Abstract
The present invention relates to a tetracycline group antibiotic ELIA kit, belonging to the field of enzymoimmunoanlysis technology. Said invention adopts the enzymoimmunoadsorption, protein coupling and biochemical preparation technique to prepare the invented detection kit. Its key technique lies in that after the tetracycline group is renovated, it is coupled with protein BSA by using succinyl oxide as bridge to prepare coating antigen, the tetracgcline group is coupled with protein BSA by using glutaraldehyde as bridge to synthesize immunoantigen, then the enzymoimmunoadsorption indirect competition method can be used to detect tetracycline group. Its detection range is 1950ng/g-1.90ng/g; its detection time only requires 4 hr.
Description
Technical field
A kind of Cyclomycin family antibiotic enzyme-linked immunoassay reagent kit the invention belongs to the EIA enzyme immunoassay technical field.Be applied to animal-derived food the detection of Cyclomycin family antibiotic content in esp meat series products, aquatic products, bee product and the dairy produce.
Background technology
Cyclomycin family antibiotic comprises aureomycin, terramycin and tetracycline etc., can be used as the veterinary drug of prevention and treatment livestock and poultry, or as the feed addictive that promotes growth of animals or poultry, is used for the treatment of multiple fish disease in culture fishery; In dairy industry, be used to improve milk production of cow; In apiculture, be used to prevent the infectious diseases of honeybee.But excessive use unavoidably makes associated antibiotics such as parent, metabolic product residue in the muscle, egg, milk, organs and tissues of food source property animal, influences health by biologic chain.Carrying out food security in the works, in view of antibiotic effect and harm and deposit, the residue problem of Cyclomycin family antibiotic especially receives publicity in the animal food, and various countries have formulated high residue amount standard of animal derived food herbal medicine and detection method.
At present, the residual detection method of Cyclomycin family antibiotic mainly is that microorganism suppresses method, chromatography and immunization etc. in the animal derived food.Microorganism suppresses method and immunoassay is applicable to conventional screening technique, physical-chemical process is used for identifying and is quantitative, but microorganism inhibition method lacks sensitivity and selectivity, physical-chemical process needs specialized office worker, sample pretreatment requires high, immunoassay is single-minded, sensitive, is that one of method of development potentiality is arranged most.And applying of enzyme linked immunological (ELISA) method must have the kit product of corresponding maturation.
Summary of the invention
The purpose of this invention is to provide a kind of simple in structurely, easy to operate, detect Cyclomycin family antibiotic enzyme-linked immunoassay reagent kit fast.The present invention be directed to the enzyme-linked immunologic detecting kit of the residual mensuration of Cyclomycin family antibiotic, preparation can produce artificial antigen, the envelope antigen of telracycline family antibody, the antibody of tetracycline resistance family, and certain cross reaction is arranged with Cyclomycin family antibiotic, related reagent is assembled into the kit that can directly use, be applied to animal-derived food, the detection of Cyclomycin family antibiotic content in esp meat series products, aquatic products, bee product and the dairy produce.
The present invention comprehensively adopts technology such as enzyme linked immunological absorption, protein coupling and biological chemistry preparation to prepare the Cyclomycin family antibiotic residual enzyme-linked immunologic detection reagent kit.Tetracycline and carrier protein couplet are prepared into artificial immunity antigen and envelope antigen; Prepare the antibody of tetracycline resistance family with artificial immunizing antigen immune animal; Telracycline family envelope antigen bag is adsorbed on the solid phase carrier; And the reagent that will detect usefulness is mixed with the reagent that can directly use.During use with standard items or sample with antibody mixes the back and the antigenic competition on the solid phase carrier combines, free antigen antibody complex is removed in washing, be combined in that antigen and enzyme mark antiantibody combines on the solid phase carrier, measure with zymolyte, in conjunction with the enzyme labeling thing colourless developer is converted into blue product.Make color change yellow into after adding reaction terminating liquid by indigo plant.Measure at the 450nm place with microplate reader, the telracycline family concentration in absorbing light intensity and the sample is inversely proportional to.
Technical scheme of the present invention: this detection kit by 1. standard solution reagent bottles, 2. enzyme mark antiantibody solution reagent bottle, 3. the antibody-solutions reagent bottle, 4. the substrate solution reagent bottle, 5. the chromogenic reagent solution reagent bottle, 6. the stop buffer reagent bottle, 7. the cleansing solution reagent bottle, 8. ELISA Plate and 9. ELISA Plate supports, 10. box body is formed.Reagent bottle 1-7 and ELISA Plate and ELISA Plate support are installed in the box body.The envelope antigen of coated elisa plate, the artificial immunity antigen of preparation antibody is by telracycline family and the protein-coupled synthetic of BSA.
ELISA Plate (8) is to wrap by the polystyrene micro-reaction plate of telracycline family antigen, and 24 holes or 48 holes or 96 holes are arranged.
The preparation of agents useful for same and the bag of ELISA Plate are:
A) preparation of standard solution: accurately take by weighing standard hydrochloric acid telracycline family 1mg, be accurate to 0.00002g, be made into 0.5mg/mL standard solution mother liquor, be filled in the reagent bottle (1), during use, be diluted to required concentration (10-1000ng/mL) with PBS (phosphate buffer).
B) enzyme mark antiantibody solution preparation: horseradish peroxidase-goat anti-rabbit igg stoste, be filled in the reagent bottle (2), during use with cleansing solution by being mixed with working concentration at 1: 4000;
C) antibody-solutions preparation: prepare resulting polyclone or monoclonal antibody with artificial immunizing antigen immune animal, the telracycline family antibody of gained was diluted to working concentration 1: 4000 with phosphate buffer, be filled into reagent bottle (3).
D) substrate solution preparation: use sodium acetate-citrate buffer solution of 0.1mol/L pH5.0, be mixed with mass concentration 0.3%H again
2O
2Make stock solution, add 14 μ L storing solutions in every 1ml damping fluid, be filled into reagent bottle (4).
E) chromogenic reagent solution preparation: be mixed with the tetramethyl biphenyl amine aqueous solution of 10mg/mL with acetone, be mixed with the tetramethyl biphenyl amine aqueous solution of 0.2mg/mL, be filled into reagent bottle (5) with sodium acetate-citrate buffer solution of 0.1mol/LpH5.0.
F) stop buffer preparation: 2mol/L H
2SO
4Solution is filled into reagent bottle (6).
G) cleansing solution preparation (PBST): contain the phosphate buffer of 0.05% polysorbas20, phosphate buffer (PBS): contain the sodium chloride of 0.15mol/L in the 0.01mol/L pH7.5 phosphate buffer, be filled into reagent bottle (7).
H) the bag quilt of ELISA Plate: the polystyrene micro-reaction plate of envelope antigen, get the artificial envelope antigen 150-200 of telracycline family μ L, add in the reaction plate hole, 4 ℃ of refrigerator overnight are poured out liquid in the hole, with cleansing solution (7) washing 3-5 time, ELISA Plate is upside down on the thieving paper pats, blot, add 200 μ L 1%BSA PBS confining liquids in the ELISA Plate aperture of envelope antigen, 37 ℃ of constant temperature are cultivated 1h, wash with cleansing solution (7), repeat 3 times, blot, encapsulation with thieving paper.
Being prepared as follows of used envelope antigen:
Telracycline family is transformed into 4-hydrazono--4-dedimethylamino tetracycline family, is bridge and carrier protein BSA coupling by succinic anhydride again, reaction equation such as figure below:
The preparation of used immunizing antigen:
Telracycline family is bridge and carrier protein BSA coupling by glutaraldehyde, and reaction equation is as follows:
Being prepared as follows of used telracycline family antibody;
Above-mentioned artificial immunity antigen-immunized animal is prepared polyclone or monoclonal antibody.
Polyclonal Antibody Preparation
The male New Zealand of body weight 1.5-2kg clinical health exempts from, with the telracycline family artificial antigen is immunogene, dosage immunity by body weight 500 μ g/kg, first with the subcutaneous or intracutaneous multi-point injection of immunogene that contains the Fu Shi Freund's complete adjuvant, after this use the immunogene booster immunization that contains freund 's incomplete adjuvant around every interval, before and after totally four times, back 10 days of last immunity, slaughter, after the blood sampling, separation of serum refrigeration standby.
MONOCLONAL ANTIBODIES SPECIFIC FOR
With the telracycline family artificial antigen is immunogene, immunity BALB/C small white mouse, immunizing dose 40 μ g, the immunogene first immunisation that contains the Fu Shi Freund's complete adjuvant, later on every around, with containing the immunogene booster immunization of freund 's incomplete adjuvant, front and back totally four times, back 10 days of last immunity, blood sampling, extracting spleen cell.Get splenocyte of immune small white mouse and the myeloma cell of small white mouse and hybridize fusion, the preparation hybridoma.Adopt limiting dilution assay screening hybridoma, the hybridoma cell strain of screening energy stably excreting telracycline family monoclonal antibody.The production of monoclonal antibody and purifying, mouse peritoneal injection hybridoma is gathered ascites, and through the ammonium sulfate precipitation purifying, packing is stand-by.
Detection method
The detection method of Cyclomycin family antibiotic detection kit of the present invention is with in standard solution or sample solution and the suitable telracycline family antibody adding ELISA Plate aperture that dilutes, blank and negative control hole are set simultaneously, 25-37 ℃ of constant temperature is cultivated 0.5-1h, pour out liquid in the hole, repeat with cleansing solution washing 2-5 time, ELISA Plate is upside down on the thieving paper pats; Add suitable dilution enzyme mark antiantibody solution in the ELISA Plate aperture, 25-37 ℃ of constant temperature is cultivated 0.5-1h, repeats to wash 3-5 time with cleansing solution, blots; Add substrate solution and chromophoric solution in ELISA Plate aperture to be measured, 25-37 ℃ of constant temperature is cultivated 15min, adds stop buffer again, immediately displaing yellow; Measure absorption value A at wavelength 450m place with microplate reader, return to zero as blank with the aperture that does not add antibody.Measuring the absorption value A in serial telracycline family standard solution hole, is that the corresponding telracycline family concentration of ordinate log value is a horizontal ordinate with the absorption value of each concentration of tetracycline, on semilogarithmic paper or with EXCEL drawing standard curve map.With the absorption value of testing sample solution, on typical curve, find corresponding tetracycline concentration, converse the content of tetracycline in the sample again.
Beneficial effect of the present invention
Prepared enzyme linked immunological quick detection kit has easy, characteristics fast and accurately.And be applicable to the detection of Cyclomycin family antibiotic in dairy produce, bee product, aquatic products, the meat products.Its sensing range is between 1950ng/g-1.90ng/g, and only need 4 hours detection time.This method average recovery rate 80%-120%, error is less than 8% in batch, and error is less than 10% between batch.
Description of drawings
Fig. 1 Cyclomycin family antibiotic detection kit synoptic diagram.
1, standard solution reagent bottle, 2, enzyme mark antiantibody solution reagent bottle, 3, the antibody-solutions reagent bottle, 4, the substrate solution reagent bottle, 5, chromogenic reagent solution reagent bottle, 6, the stop buffer reagent bottle, 7, the cleansing solution reagent bottle, 8, ELISA Plate, 9, the ELISA Plate support, 10, box body.
Fig. 2 ELISA Plate synoptic diagram.
Embodiment
Embodiment 1
The preparation of 1 envelope antigen
Weigh quadracycline 1 gram and be dissolved in the 50mL water, add 2.7 gram chlorosuccinimides, react after 30 minutes, filter, wash semifinished product with water.Semifinished product is handled with 150mL water and ether (volume ratio 1: 1), abandons water, and evaporate to dryness ether phase washes with water again, filters, and promptly gets pure product (about 5.6 grams), for transforming tetracycline.
Get tetracycline and the 6mg succinic anhydride that 20mg transforms and be dissolved in the 2mL dioxane, put 37 ℃ of reaction 2h, put 4 ℃ and spend the night, this is 1. liquid.Other gets 35mg ovalbumin (OVA) and is dissolved in 2mL water, adds the 1mL dioxane again, and this is 2. liquid.Get 1-ethyl-3-(3-dimethylamine propyl) carbodiimide hydrochloride (EDC) 125mg again and be dissolved in the 2mL water, this is 3. liquid, gets 3. liquid 1mL and slowly adds 1. and stir 0.5h in the liquid, and 2. this reactant liquor is slowly splashed in the liquid, stirs 4h, adds 3. liquid 0.5mL again.After continue stirring 4h, add 3. liquid 0.5mL again, stirring is spent the night, and reclaim after to distill water dialysis next day.
The preparation of 2 immunizing antigens
Take by weighing quadracycline 170mg and be dissolved in 2ml methyl alcohol, slowly add in the 0.1mol/mL pH4.0 acetic acid 100ml damping fluid, add 8ml 25% glutaraldehyde solution, 20mg BSA more successively, stirring at room 5 hours.The SephadexG-75 chromatographic column (1 * 22cm) that reactant liquor is crossed by 0.01mol/mL pH4.0 acetate buffer solution balance, carry out wash-out with same damping fluid, fraction collector is collected in the eluent that there is and deposits the peak at 280nm and 356nm wavelength place, absorbs to concentrate with Macrogol 6000 to obtain artificial immunity antigen tetracycline-glutaraldehyde-BSA.
3 telracycline family Polyclonal Antibody Preparation
The male New Zealand of body weight 1.5-2kg clinical health exempts from, with tetracycline-glutaraldehyde-BSA is immunogene, by the dosage of body weight 500 μ g/kg immunity four times, for the first time with the subcutaneous or intracutaneous multi-point injection of immunogene that contains 1.5mL Fu Shi Freund's complete adjuvant, around after this every interval, with the immunogene booster immunization that contains the 1.5mL freund 's incomplete adjuvant, front and back totally four times, the growth and decline situation of antibody is measured in blood sampling behind each booster immunization, once immune back 10 days of end, slaughter, after the blood sampling, separation of serum refrigeration standby.
The preparation of 4 kits
A) the polystyrene micro-reaction plate of envelope antigen: get the antigenic solution 200 μ L of 1-10 μ g/ml concentration, join in the ELISA Plate aperture, shake up, put 4 ℃ of refrigerator overnight.Pour out liquid in the hole,, repeat 3-5 time, ELISA Plate is upside down in pats on the thieving paper, blot with the washing of PBST cleansing solution.Add 200 μ L 1%BSA PBS confining liquids in the ELISA Plate aperture of envelope antigen, 37 ℃ of constant temperature are cultivated 1h, with the cleansing solution washing, repeat 3 times, blot with thieving paper.Encapsulation.
B) preparation of standard solution: accurately take by weighing standard quadracycline 1mg (being accurate to 0.00002g), be made into the 0.5mg/mL mother liquor, be diluted to required concentration (10-1000ng/mL) with PBS.Can is a reagent 1.
C) enzyme mark antiantibody solution: horseradish peroxidase-goat anti-rabbit igg stoste.Can is a reagent 2.
D) tetracycline antibody-solutions: tetracycline resistance antibody with phosphate buffer by being diluted to working concentration at 1: 4000.Can is a reagent 3.
E) substrate solution: the sodium acetate-citrate buffer solution with 0.1mol/L pH5.0 is mixed with 0.3%H earlier
2O
2Stock solution adds 14 μ L stock solutions in every 1mL damping fluid.Can is a reagent 4.
F) chromophoric solution:, be mixed with the tetramethyl biphenyl amine aqueous solution of 0.2mg/mL with sodium acetate-citrate buffer solution of 0.1mol/L pH5.0 earlier with the tetramethyl biphenyl amine aqueous solution of acetone preparation 10mg/mL.Can is a reagent 5.
G) stop buffer: 2mol/L H
2SO
4Solution.Can is a reagent 6.
H) cleansing solution (PBST): the phosphate buffer that contains 0.05% polysorbas20.Phosphate buffer (PBS): the sodium chloride that contains 0.15mol/L in the 0.01mol/L pH7.5 phosphate buffer.Can is a reagent 7.
Above-mentioned reagent bottle, ELISA Plate and ELISA Plate support are packed in the box, form the tetracycline enzyme-linked immunologic detecting kit.
Antigen: use occrycetin standard items, antibody instead: use the terramycin antibody-solutions instead.ELISA Plate is used terramycin envelope antigen bag quilt instead, and the configuration of all the other reagent bottles is the terramycin enzyme-linked immunologic detecting kit with embodiment 1.
Antigen: use aureomycin hydrochloride standard items, antibody instead: use chlortetracycline antibody solution instead.ELISA Plate is used aureomycin envelope antigen bag quilt instead, and the configuration of all the other reagent bottles is the aureomycin enzyme-linked immunologic detecting kit with embodiment 1.
1 sample preparation
A) the dairy products skimmed milk can be that 1: 10 dilution back is directly measured with skimmed milk: PBS with PBS, and the whole milk then should measure after PBS was with dilution in 1: 10 behind the centrifugal degrease again.
B) honey take by weighing honey 10g with the dissolving of 10% ethanol-PBST and constant volume to 25mL, stir and filter, get filtrate mensuration.
C) royal jelly takes by weighing mixed uniform royal jelly 200mg, places the centrifuge tube of band scale, adds pH8.0PBS to 2mL scale place and dilution evenly, and the centrifuging and taking supernatant is measured.
D) meat products takes by weighing sample 5g with a small amount of 10% ethanol-PBST, puts in the glass grinding device, grinds to form even pasty state, wash out with 10% ethanol-PBST then, and constant volume, through the refrigerated centrifuge degrease, is got supernatant and measured after stirring to 25mL.
E) aquatic products take by weighing sample 5g in glass homogenizer, add a small amount of 2.0% casein-PBST, behind the homogeneous with its constant volume to 20mL, get supernatant after centrifugal and detect.
2 determination steps
A) prepare
Before analyzing with all reagent balances to room temperature; On request relevant reagent is diluted to working concentration; Immediately all reagent are put back to 4 ℃～8 ℃ refrigerators after the analysis; In all were cultivated, lucifuge covered the microwell plate lid.
B) reagent configuration
The dilution of ELIAS secondary antibody (reagent 2): get ELIAS secondary antibody 250 μ L before the use and be diluted to use liquid with washing agent (reagent 7).
C) location
Set limit the quantity of method (seeing Table 1) and sizing technique (seeing Table 2) as required.The ELISA Plate micropore of getting sufficient amount places on the ELISA Plate support position of record standard product hole and sample aperture.Concentration when limiting the quantity of method in the control gauge orifice number value/extension rate that exceeds, and make it concentration in the 10-1000mg/mL measurement range by regulating extension rate.
Limit the quantity of method micropore location of table 1
| Zero standard hole number | Control criterion hole number | Sample well number | |||||
| 1 | 2 | 3 | 4 | 5 | 6 | 7 | 8 |
Table 2 sizing technique micropore location
| Gauge orifice concentration μ g/L | Sample well number | ||||||||||
| a | b | c | d | e | f | 1 | 2 | 3 | 4 | 5 | 6 |
D) immune response
In every hole, add reagent successively, add 50 μ L standard solution or sample extract to corresponding micropore; Add 50 μ L antibody-solutions again in each micropore.Shake up, ELISA Plate is placed in the polybag, put the dark place, room temperature (25 ℃～37 ℃) reaction 1 hour.Liquid pouring in the micropore in the pond, is inverted the micropore support, on clean paper handkerchief, pats, remove all residual liquid, add the about 250 μ L of cleansing solution and in each micropore, wash plate, evacuation of liquid again, repeated washing 4 times.In every hole, add 100 μ L ELIAS secondary antibody solution successively in each micropore.Shake up, ELISA Plate is placed in the polybag, put the dark place, room temperature (20 ℃～30 ℃) reaction 1 hour.Wash by above-mentioned washing methods.
E) chromogenic assay
Every hole adds substrate solution and each 50 μ uL (being equivalent to one) of chromogenic reagent solution respectively, fully shakes up, and puts the dark place, room temperature (25 ℃～37 ℃) reaction 15min.(drip to squeeze earlier before the solution at every turn and splash into micropore again after going 2-3 to drip.) add 50 μ L (being equivalent to) stop buffer and in every hole, shake up.At the 450nm place, is blank zeroing with the air with microplate reader, measures absorption value.Reading in 60min.
3 results calculate and statement
A) method of limiting the quantity of
If the absorption value of sample aperture is less than the absorption value of gauge orifice, i.e. A
450nmSample aperture＜A
450nmGauge orifice surpasses the value of limiting the quantity of, and is positive.If the absorption value in the absorption value overgauge hole of sample aperture, i.e. A
450nmSample aperture＞A
450nmGauge orifice, then, negative less than the set value of limiting the quantity of.
B) sizing technique
The drafting of typical curve: the semilog coordinate of the corresponding telracycline family of the absorption value of the standard items of surveying (μ g/L) is made canonical plotting, and curve within the specific limits should be linear.According to the percentage absorption value of sample,, check in corresponding concentration by typical curve.By the telracycline family content in formula (1) the calculating sample.
Telracycline family content X in the sample, unit are every kilogram of microgram (μ g/kg), are calculated as follows;
In the formula:
C-checks in telracycline family concentration the corresponding extract from typical curve, and unit is every liter of microgram (μ g/L);
V-sample extracting liquid volume, unit are milliliter (mL);
N-sample extension rate;
The m-sample mass, unit is gram (g).
Result of calculation is represented a position effective digital behind the radix point.
Claims (5)
1. Cyclomycin family antibiotic enzyme-linked immunoassay reagent kit, it is characterized in that by standard solution reagent bottle (1), enzyme mark antiantibody solution reagent bottle (2), antibody-solutions reagent bottle (3), substrate solution reagent bottle (4), chromogenic reagent solution reagent bottle (5), stop buffer reagent bottle (6), cleansing solution reagent bottle (7), ELISA Plate (8) and ELISA Plate support (9), box body (10) is formed, reagent bottle (1), (2), (3), (4), (5), (6), (7) and ELISA Plate (8) and ELISA Plate support (9) be installed in the box body (10), the envelope antigen of coated elisa plate, the artificial immunity antigen of preparation antibody is by telracycline family and the protein-coupled synthetic of BSA;
A) standard solution preparation: accurately take by weighing standard hydrochloric acid telracycline family 1mg, be accurate to 0.00002g, be made into 0.5mg/mL standard solution mother liquor, be filled into reagent bottle (1), during use, be diluted to the series standard product solution of required concentration 10-1000ng/mL with phosphate buffer;
B) enzyme mark antiantibody solution preparation: enzyme mark antiantibody is horseradish peroxidase-goat anti-rabbit igg stoste, is filled into reagent bottle (2), during use with cleansing solution by being mixed with working concentration at 1: 4000;
C) antibody-solutions preparation: prepare resulting polyclone or monoclonal antibody with artificial immunizing antigen immune animal, the telracycline family antibody of gained was diluted to working concentration 1: 4000 with phosphate buffer, be filled into reagent bottle (3);
D) substrate solution preparation: use sodium acetate-citrate buffer solution of 0.1mol/L pH5.0, prepare mass concentration 0.3%H again
2O
2Make stock solution, add 14 μ L stock solutions in every 1ml damping fluid, be filled into reagent bottle (4);
E) chromogenic reagent solution preparation: be mixed with the tetramethyl biphenyl amine aqueous solution of 10mg/mL with acetone, be mixed with the tetramethyl biphenyl amine aqueous solution of 0.2mg/mL, be filled into reagent bottle (5) with sodium acetate-citrate buffer solution of 0.1mol/LpH5.0;
F) stop buffer preparation: 2mol/L H
2SO
4Solution is filled into reagent bottle (6);
G) cleansing solution preparation: contain the phosphate buffer of 0.05% polysorbas20, contain the sodium chloride of 0.15mol/L in the 0.01mol/L pH7.5 phosphate buffer, be filled into reagent bottle (7);
H) the bag quilt of ELISA Plate: get the artificial envelope antigen 150-200 of telracycline family μ L, add in the reaction plate hole, 4 ℃ of refrigerator overnight are poured out liquid in the hole, with cleansing solution (7) washing 3-5 time, ELISA Plate is upside down on the thieving paper pats, blot in the ELISA Plate aperture of envelope antigen and add 200 μ L 1%BSA PBS confining liquids, 37 ℃ of constant temperature are cultivated 1h, wash with cleansing solution (7), repeat 3 times, blot, encapsulation with thieving paper.
2. detection kit according to claim 1 is characterized in that ELISA Plate (8) is to wrap by the polystyrene micro-reaction plate of telracycline family antigen, has 24 holes or 48 holes or 96 holes.
3. detection kit according to claim 1 is characterized in that the preparation of envelope antigen: it is bridge and carrier protein BSA coupling by succinic anhydride again that telracycline family is transformed into 4-hydrazono--4-dedimethylamino tetracycline family.
4. detection kit according to claim 1 is characterized in that the preparation of artificial immunity antigen: telracycline family is bridge and carrier protein BSA coupling by glutaraldehyde.
5. detection kit according to claim 1 is characterized in that described Cyclomycin family antibiotic is tetracycline or aureomycin or terramycin.
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|---|---|---|---|
| CN 200610037948 CN1811454A (en) | 2006-01-19 | 2006-01-19 | Cyclomycin family antibiotic enzyme-linked immunoassay reagent kit |
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN 200610037948 CN1811454A (en) | 2006-01-19 | 2006-01-19 | Cyclomycin family antibiotic enzyme-linked immunoassay reagent kit |
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| US7858601B2 (en) | 2004-10-25 | 2010-12-28 | Paratek Pharmaceuticals, Inc. | 4-substituted tetracyclines and methods of use thereof |
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| US7858601B2 (en) | 2004-10-25 | 2010-12-28 | Paratek Pharmaceuticals, Inc. | 4-substituted tetracyclines and methods of use thereof |
| CN101315375B (en) * | 2008-06-30 | 2012-08-08 | 江南大学 | ELISA detection reagent kit suitable for Nitrazepam relict analysis |
| CN102207505A (en) * | 2010-03-29 | 2011-10-05 | 上海友科生物科技有限公司 | Method for in vitro detection of zinc-alpha2-glycoprotein, and kit thereof |
| CN102175850A (en) * | 2010-12-30 | 2011-09-07 | 北京肿瘤医院 | ELISA (enzyme linked immunosorbent assay) kit for detecting endothelial cell specific molecule-1 (ESM-1) of tumour marker |
| CN102288753A (en) * | 2011-05-10 | 2011-12-21 | 重庆市科学技术研究院 | Quadruple colloidal gold immunochromatography testing strip for rapid detection of residual tetracycline, chlortetracycline, oxytetracycline and doxycycline and preparation method of same |
| CN102818888A (en) * | 2012-07-31 | 2012-12-12 | 白仲虎 | Test box |
| CN103421108A (en) * | 2013-08-13 | 2013-12-04 | 山东绿都生物科技有限公司 | Method of synthesizing tetracycline coupled antigen |
| CN103421108B (en) * | 2013-08-13 | 2015-01-07 | 山东绿都生物科技有限公司 | Method of synthesizing tetracycline coupled antigen |
| CN105001112A (en) * | 2015-06-30 | 2015-10-28 | 浦城正大生化有限公司 | Water soluble chlorotetracycline succinic acid monoester salt, and preparation method thereof |
| CN107543928A (en) * | 2017-08-24 | 2018-01-05 | 太原瑞盛生物科技有限公司 | A kind of chemiluminescence detection kit of aureomycin and preparation method thereof |
| CN109613159A (en) * | 2018-12-25 | 2019-04-12 | 南京祥中生物科技有限公司 | Method that is a kind of while detecting Tetracyclines, lincomycin, Florfenicol antibiotic residual quantity |
| CN109574868A (en) * | 2018-12-28 | 2019-04-05 | 天津阿尔塔科技有限公司 | A kind of preparation method of Tetracyclines and its deuterated internal standard compound of epimer |
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