A simple assay system for infection-enhancing and -neutralizing antibodies to dengue type 2 virus using layers of semi-adherent K562 cells.

A simple alternative to the dengue antibody-dependent enhancement (ADE) assay was established. The new assay method utilizes cells attached to microplate wells, thereby eliminating cumbersome procedures typical of the conventional ADE assay that utilizes suspension cells. Semi-adherent K562 cells bearing the Fc-gamma receptor (Fc gammaR) were cultured on poly-L-lysine-coated plates. The procedure consisted of (i) preparation of a virus-antibody-cell mixture in wells, (ii) cultivation at 37 degrees C for 24 h and (iii) fixation and immunostaining to count infected cells. Using monoclonal antibodies against dengue type 2 virus, the new system correlated with three conventional systems. Additionally, K562 cells were employed in a neutralization test. For this purpose, the virus-antibody mixture was incubated at 37 degrees C for 2 h prior to the addition of cells. As expected, K562 cells provided lower neutralizing antibody titers than did a conventional neutralization test using Vero cells, which do not have Fc gammaR, in monoclonals showing both neutralizing and enhancing activities. Since antibodies are present in polyclonal form in circulation, neutralization tests using K562 cells are considered to reveal a more accurate in vivo status than those using Vero cells. Human sera, positive for dengue virus antibodies, showed neutralizing and enhancing activities in a dose-dependent manner. 2009 Elsevier B.V. All rights reserved.