An in vitro microassay for lymphotoxin using microculture plates and the multiple automated sample harvester

Abstract

An in vitro microassat for lymphotoxin (LT) using mouse L-929 fibroblasts as target cells grown in the wells of microculture plates is described. The microassay uses 1000 targets cells and 50 μl of LT-containing fluids per well. Destruction or inhibition of target cell growth in the presence of LT-containing fluids is measured by their ability to incorporated 3H-methyl-thymidine. Radioactively labelled cells are harvested onto glass fiber filter discs using a multiple automated sample harvester and are subsequently counted in a liquid scintillation spectrometer. Inhibitory or destructive activity of LT-containing fluids can be conveniently represented as the dilution which inhibits target cell growth by 50 percent (ID 50). The advantages and limitations of the microassay are shown using LT-containing supernatant fluids from human lymphoblast cell lines and PHA-P- and PHA-M-stimulated human lymphocytes.