Microdroplet Assay of Human Serum Cytotoxins

Abstract

ATTEMPTS to define leucocyte groups in man serologically have been hampered by the scarcity of immune human antisera and the capriciousness of the leuco-agglutination reaction¹. The method recorded here for assaying lymphocyte cytotoxins in a microscale was developed to circumvent these difficulties. Its extreme sensitivity permits performance of 1,000 or more tests with 1 ml. of antiserum. Furthermore, lymphocytes obtained from one finger-prick sample of blood are sufficient for 100 separate tests. The basic innovation in handling these small quantities of serum and cells and in gaining a greater than ten-fold increase in sensitivity over present-day methods is the performance of the reaction in microdroplets submerged under oil. This permits reduction in numbers of target cells to as few as 500 cells in contrast to the 50,000 or so cells required for previously described cytotoxicity tests^2–4. Since the sensitivity of cytotoxicity is inversely proportional to the number of cells used^2,3, an increase of sensitivity in the range of 10–100 times may be expected. Unlike the usual de Fonbrune oil chamber employed in micromanipulation, use of a ‘dish’ with a cover-glass bottom facilitated rapid addition of reagents and cells with a microsyringe.