Determination of antibody response to influenza virus surface glycoproteins by kinetic enzyme-linked immunosorbent assay

Abstract

We modified an existing enzyme-linked immunosorbent assay (ELISA) to be able to use new spectrophotometers which can measure the rate of color development in microtiter wells. This new kinetic-based ELISA (KELISA) required only a single dilution of specimen rather than the multiple dilutions required with endpoint ELISA. In addition, 10- to 100-fold-less specimen was required to perform the KELISA than the ELISA. The level of serum or nasal wash antibody against surface glycoproteins of influenza A or influenza B viruses determined by KELISA was reproducible and correlated highly with the results of endpoint ELISA or hemagglutination inhibition tests. The difference between the KELISA rates, which indicated than an antibody response to infection had occurred, was defined and was analogous to a 2.2-fold rise in titer for serum and a 3.4-fold rise in titer for nasal wash determined by endpoint ELISA. The KELISA was similar to endpoint ELISAs in its ability to detect rises in antibody level in paired serum or nasal wash specimens obtained from volunteers who received live attenuated influenza A reassortant virus vaccines. By eliminating the need for multiple dilutions, the use of KELISA offers the advantage of increasing the number of assays that can be performed by the same personnel compared with endpoint ELISA, while it maintains sensitivity and specificity.