[Measurement method] ELISA method
[Methodological principle] The purified EB virus antigen is pre-coated on the microplate. If EBVAb-IgA is present in the sample to be tested, it can be combined with the pre-coated EB-VAg on the microplate, and then the enzyme is added. The reagent binds to the antigen-antibody complex, and a color reaction occurs under the action of the color developer, and the color depth is positively correlated with the antibody content in the sample.
[Preparation of specimens] 2ml of venous blood, not anticoagulation.
[Reagents]
1. Microplate
2. Sample diluent
3. Enzyme conjugate
4. Negative control
5. Positive control
6. Detergent
7. TMB coloring solution
8. Stop solution
[Instrument equipment] Microplate reader or fully automatic enzyme-linked immunosorbent detector.
[Measurement step]
1. The serum to be tested was numbered and diluted 1:100 with the sample dilution.
2. The microporous strips of the corresponding number of holes were taken out according to the amount, and the remaining portions were stored at 4 ° C after sealing.
3. Add diluted serum to be tested, negative control, positive control each 100t. Tl to the reaction well, the negative control, the positive control set up the reaming, set a blank control 1 well (no blank added to the blank), shake and mix, incubate for 1 h in 37C environment.
4. Discard the liquid in the well, wash it 5 times with the washing solution, and pat dry.
5. The enzyme marker 100fA was added to each well, mixed, and bathed in a water bath at 37 ° C for 1 h.
6. Discard the liquid in the well, wash it 5 times with the washing solution, and pat dry.
7. Two drops of substrate A and substrate B were added to each well, and the mixture was allowed to stand at room temperature for 10 min, and 50 μl of the stop solution was added to each well to terminate the reaction.
8. The blank control (only 200 g1 of distilled water) was zeroed, and the OD value of each well was read at a wavelength of 450 nm. The sample OD was greater than or equal to 2 times of the negative control well, and the sample was positive.
[normal reference value] EBVAb-IgA negative
[Precautions]
1. The kit should be equilibrated for 30 min at room temperature before use.
2. The kit should be stored at 2～8°C in the dark and should be used up as soon as possible after activation.
3. Different batches of reagent components cannot be mixed.
4. After the experiment, the reagents and serum samples should be regarded as infectious substances.
5. When dropping with a dropper, the dropper should be vertical, and the force and speed should be uniform; mix well after loading.
[Clinical significance] EBVAb-IgM positive is an indicator of recent infection of EBV, which is common in infectious mononucleosis. The antibody turns to yang soon after the patient has been infected, but usually has a shorter duration.
EBVAb-IgA is positive in nasopharyngeal carcinoma and can be used for observation and prognosis of nasopharyngeal carcinoma. It can also be seen in lung cancer, thyroid cancer, and chronic nasopharyngeal inflammation, but the positive rate is generally not high. Normal people are also partially positive, with a positive rate of about 3.4%.
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