Assay for Plaque-Forming Cells

Publisher Summary This chapter discusses assay for plaque-forming cells. Plaque assay includes lymphoid cells that are mixed with a suspension of red blood cells and immobilized in a gel or in a liquid medium enclosed in a sealed chamber. The specific antibody synthesized by some of the lymphoid cells is released and diffuses from the central cell; antibody is trapped by antigen in the areas immediately surrounding the plaque-forming cell (PFC). In the presence of complement, plaques will appear as a consequence of lysis of sensitized red cells. The fresh red cells are considerably less susceptible to lysis than aged red cells, and one should use at least 1-week-old red blood cells, stored in Alsever's solution, as indicator cells. If PFCs for nonhemolytic classes of antibody have to be revealed, developing serum is added to the samples. The optimal dilution of developing serum to be used must be determined empirically. Basically two approaches can be used: (1) the developing serum is added together with complement and the total number of direct and indirect plaques is counted and (2) complement is added alone, the mixture is incubated, direct plaques are counted, the complement is poured off, and finally developing serum and fresh complement are added and incubated and newly developed indirect plaques are counted.