BT Lab Human Papillomavirus Antibody, IgM Elisa Kit

Assay Principle 
This kit is based on a Qualitative reverse phase enzyme immunoassay technique. The microtiter plate has been pre-coated with a target antigen. Positive/Negative Controls or samples are added to the wells and incubate. Antibodies in the samples bind to the antigen on the plate. Unbound antibody is washed away during a washing step. A Horseradish Peroxidase (HRP) conjugated detection antibody is then added and incubate. Unbound HRP is washed away during a washing step. TMB substrate is then added and color develops. The reaction is stopped by addition of acidic stop solution and color changes into yellow that can be measured at 450 nm. The OD of an unknown sample can then be compared to the OD of the positive and negative controls in order to determine the presence of HPV Ab IgM.

Summary
1. Prepare all reagents, samples and controls and set a blank control with no solution.
2. Add 50ul positive control or negative control to each well.
3. Dilute 10ul sample with 40ul sample diluent and add this diluted sample to each well.
4. Incubate for 30 minutes at 37°C.
5. Aspirate and wash 5 times.
6. Add HRP to each well and incubate for 30 minutes at 37°C.
7. Aspirate and wash 5 times.
8. Add substrate solution A and substrate solution B and incubate in the dark for 10 minutes at 37°C.
9. Add 50ul stop solution.
10. Read the OD value within 15 minutes at 450nm.

Calculation of Result 
Average the reading from duplicate or triplicate samples. For calculation the valence of Human Papillomavirus Antibody, IgM ELISA Kit, compare the sample well with control.