Objective: Tetanus is a vaccine preventable disease that is seen mostly in the developing countries. In order to evaluate the efficacy of vaccination programmes or to evaluate the immune status of the population, anti-tetanus antibody levels must be investigated by serological studies. ELISA test is a simple, rapid and economical test for the detection of tetanus antibody titers. However, the results may be affected by cross reactive antibodies or low levels of antibodies may remain undetected. Our aim was to improve an in-house ELISA kit by blocking the cross reacting antibodies and enable the detection of low levels of antibodies.

Materials and Methods: The selected 322 serum samples were tested for the presence of anti-tetanus antibodies by an in-house ELISA method, named as ELISA 1. The sera were grouped according to the levels detected by ELISA 1 test as ≤0.01 IU/ml, 0.01-0.1 IU/ml, 0.1-1 IU/ml, 1-10 IU/ml, >10 IU/ml. A total of 52 sera were tested for tetanus antibody levels by the gold standard mouse neutralization (NT) test. Sera with antibody levels below 0.01 IU/ml (n=25) and the sera randomly selected from the other groups (n=27) were included in the neutralisation test group. Samples tested with neutralization were later also tested by the modified ELISA 2 method.

Results: NT and ELISA 1 test results correlated well for the sera with antibody level ≥1.0 IU/ml. However, for antibody levels below 1.0 IU/ml, ELISA 1 test results were higher than NT results. The results of ELISA 2 method was better than ELISA 1 test when NT results were considered as the gold standard. When ELISA 2 results were compared to NT results, the lowest reliable detection limit was decreased to 0.5 IU/ml.

Conclusion: The lowest antibody detection limit for ELISA 2 method was 0.5 IU/ml. In order to detect antibody levels 0.01 IU/ml and below, further studies should be performed to improve the in-house ELISA method.