Highly pathogenic avian influenza H5 antibody detection kit

Highly pathogenic avian influenza H5 antibody detection kit (enzyme-linked immunoassay)

Product name

Generic Name: Universal Avian Influenza Virus Antibody Test Kit (Enzyme-Linked Immunoassay)

Packaging specifications

96T ×1/box

Detection principle

This kit consists of a pre-coated enzyme-labeled plate, enzyme labels, and other supporting reagents coated with avian influenza H5 recombinant protein, and uses the principle of enzyme-linked immunosorbent (ELISA) to detect antibodies against avian influenza H5 virus in chicken serum and plasma samples. During the experiment, the control serum and the sample to be tested were added to the microplate labeled plate. After incubation, if the sample contained antibodies against avian influenza H5 virus, they would bind to the antigen on the plate, and after washing to remove unbound components, Enzyme markers are then added to specifically bind to antigen-antibody complexes on the microplate labeled plates; Then, wash to remove unbound enzyme markers, add TMB substrate solution to the well, and react with the enzyme-labeled conjugate to form blue products. The depth of color development is positively correlated with the specific antibody content in the sample; After adding the termination solution to terminate the reaction, the product turns yellow; By measuring the absorbance value in each reaction well at a wavelength of 450nm using an microplate reader, it can be determined whether the sample contains antibodies against avian influenza H5 virus.

Composition of the kit

Storage and expiration date

Store at 2~8°C, shelf life is 12 months.

After opening, please store the blanket board at 2~8°C away from light to avoid moisture. The usage period is 2 months.

Applicable instruments

Includes a 450nm wavelength microplate reader, 37°C constant temperature equipment, and adjustable micropipettes.

Sample preparation

1. Prepare serum from whole animal blood using conventional methods, ensuring the serum is clear, free of hemolysis, and free of contamination. Samples can be stored at 2~8°C for 1 week, and should be stored at -20°C for extended periods.

2. Prepare the corresponding number of small-scale EP tubes or serum dilution trays according to the sample quantity, and use the sample diluent to dilute the serum to be tested at a 50-fold ratio (e.g., add 4μl serum to 196μl sample dilution and mix well). Negative and positive controls do not require dilution.

3. Before use, the concentrated detergent should be returned to room temperature to dissolve the precipitate, then dilute 20 times with distilled or deionized water to make a working detergent (e.g., 19 parts distilled or deionized water plus 1 part concentrated detergent).

Inspection methods

1. Before use, place the kit at room temperature for 30 minutes to restore to room temperature.

2. Take the required amount of microplate-labeled slats, set one well each for negative and positive controls, seal unused slats as soon as possible, and store at 2~8°C.

3. Add 100μl of negative and positive controls to the negative and positive control wells; Add 100μl of the diluted sample to each well.

4. Mix well, cover with the cover film, and incubate in a 37°C incubator for 30 minutes.

5. Shake off the liquid inside the holes, add 250μl of working cleaning solution to each hole, wash evenly by shaking for 60 seconds, then discard. Repeat the wash 5 times, and after each wash, pat dry the liquid.

6. Add 100μl of enzyme labeler to each well and react at 37°C in a dark place for 30 minutes.

7. Drain the liquid inside the holes, add 250 μl of working cleaning solution per hole, wash evenly by shaking for 60 seconds, then discard. Repeat the wash 5 times, and after each wash, pat dry the liquid.

8. According to the sample quantity, mix substrate solution A and substrate solution B in equal volume proportions to prepare the substrate working liquid, ready for immediate use. Add 100 μl to each well, incubate at 37°C in a dark place for 10 minutes.

9. Add 100μl of stop solution to each well, mix well, and measure the absorbance value (OD value) of each well at 450nm.

Reference values

Under normal experimental conditions, the positive control OD versus negative control OD > 0.2.

Interpretation of test results

1. S/P = (sample OD - negative control OD) / (positive control OD - negative control OD).

2. S/P ≥ 0.4 is considered positive; 0.2< S/P < 0.4 is considered suspected infection; S/P < 0.2 is negative.

Limitations of the testing methods

1. This kit is only for preliminary assessment of whether the subject is infected with avian influenza H5 virus; please refer to clinical conditions for specific situations.

2. This kit is for research purposes only and must not be used in clinical or human experiments.

Notes

1. Gloves and work clothes must be worn during experiments, and disinfection and isolation protocols must be strictly enforced and implemented. All experimental waste must be treated as infectious agents.

2. Terrators are corrosive and should be kept away from contact with skin and clothing. If accidentally touched, rinse immediately with plenty of tap water.

3. When microplate plates are removed from refrigeration, they should be returned to room temperature before opening. Unused plates should be stored in sealed bags containing desiccant.

4. Concentrated detergents tend to crystallize at low temperatures and should be returned to room temperature for complete dissolution during use.

5. When washing, fill all holes with liquid to prevent free enzymes from occurring at the openings and preventing cleaning.

6. Samples used for testing should be kept fresh.

7. Test results must be determined based on microplate reader readings.

8. Reagent components of different batches must not be mixed.