Further attempts to characterize the normal incomplete cold antibody.

Attempts at eluting the normal incomplete cold (n.i.c.) antibody from sensitized red cells or red cell stroma, using standard methods, were unsuccessful; thus the antibody could not be detected in the eluates obtained by heating at 56° or 37° or by dissociation at acid pH. The n.i.c. antibody was partially eluted from sensitized red cells only when elution was carried out at 37° into serum instead of saline. Elution of the antibody from dextran (Sephadex G-200)—n.i.c. antibody complex formed at 0° was also achieved using a 15 per cent NaCl solution at 37°. Since it has been shown that red cells sensitized with the n.i.c. antibody are not agglutinated by anti-γG, anti-γA or anti-γM-globulin sera, an attempt at producing an antibody specifically reacting with the n.i.c. antibody was made by injecting into a rabbit the eluate obtained from zymosan—n.i.c. antibody complex. The immune serum was found to inhibit the n.i.c. antibody activity when added to normal serum and to interfere with the property of normal serum requiring the properdin system. In the present work it is also confirmed that the antibody is not associated with γG or γM-globulin; thus adult and cord sera and one serum from a patient with severe hypogammaglobulinaemia were fractionated with zone electrophoresis or DEAE-cellulose chromatography and it was found that the antibody was not present in the fractions containing the bulk of γG or γM-globulin.