The quantitation of rabies-specific antibodies. III. A comparative evaluation of modified counter immunoelectrophoresis, haemagglutination inhibition and serum neutralization titres of human sera.

The rabies-specific antibodies of 73 serum samples from vaccinated humans were determined by the modified counter immunoelectrophoresis (MCIE), and the haemagglutination inhibition test (HAI) by using the conventional serum neutralization test (SN) as a yard-stick. Both MCIE and HAI were found to be sensitive and specific for the estimation of rabies antibodies. In general, the unitages obtained by the MCIE and SN showed statistically insignificant differences (P greater than 0.05) and the correlation coefficient between the two methods was 0.697 (P less than 0.05). Although the unitage of the sera detected by HAI tests was lower by a factor of 0.155 from the unitage of SN tests, there was statistically insignificant differences between the two techniques (P greater than 0.05) with a correlation coefficient of 0.556 (P less than 0.05).

The quantitation of rabies-specific antibodies. I. Modified counter immunoelectrophoresis. A rapid and sensitive method.

Modified counter immunoelectrophoresis was standardized with respect to dilution of tissue culture antigen and indicator serum, the incubation time for neutralization and the effect of an electric current. The technique was found to be sensitive enough to detect a minimum level of antibodies (0.5 IU/ml) by using a 16 mA current per slide for 2 h, indicator serum of 15 IU/ml and the use of an antigen at a concentration of 1:35. Above all, the incubation period did not affect the neutralization of the virus. The test was also applied to the detection of rabies-specific antibody levels in 73 human sera. The test was found to be simple, quick and economical for titration of rabies antibodies.

The quantitation of rabies-specific antibodies. II. Factors affecting the sensitivity of the haemagglutination inhibition test.

Various factors affecting the HAI test for the quantitation of rabies-specific antibodies have been evaluated with a view to obtaining maximum sensitivity and reproducibility in tests using tissue culture antigens prepared in vero cells and concentrated by dialysis. Goose erythrocytes treated with proteolytic enzyme bromelian at a concentration of 0.025% were much more susceptible to HA than those that were untreated or erythrocytes treated with neuraminidase. In addition, other parameters like the use of a phosphate buffered saline (PBS) as a diluent at pH 6.2, incubation at 0-4 degrees C for 1.5-3 h were found to be most critical for achieving maximum HA activity. To remove non-specific inhibitors, serum samples were treated with aerosil, acetone in combination or alone. Of the 73 serum samples tested, removal of non-specific inhibitors by aerosil alone occurred in up to 54.79% of the samples, whereas using acetone-aerosil treatment followed by adsorption with goose erythrocytes, the inhibitors were removed in 98.67% of the samples to a level that was undetectable at the 1:4 starting dilution in the HAI test.