Methods for the purification and quantitation of IgD

Methods of isolation of IgD from D-myeloma and normal sera have been explored as has a sensitive method to quantitate low levels of IgD. DEAE-cellulose chromatography of a 30-45% saturated ammonium sulfate fraction of D-myeloma serum, yielded an IgD rich fraction in which IgG contamination did not exceed 20-25%. This was fractionated on G-200 Sephadex and IgD was eluted free of IgG in the ascending half of the first protein peak. Specific antisera to IgD were insolubilized by copolymerization and by coupling to an insoluble carrier and were used to specifically adsorb IgD from normal sera. IgD was eluted from the immunoadsorbant with solutions of high molarity. Other fractions containing IgG and IgD were purified by removing IgG vnth specific anti-IgG immunoadsorbants. IgD levels of less than 0.1 mg/ml were quantitated by a radioimmunosorbent method which depends on the competition between IgD in an unknown and 125-labelled IgD in binding to an insolubilized antiserum to IgD. Levels in excess of 0.1 mg/ml were quantitated in Cudin tubes

The organ sources(s) of the cells mediating a cellular immune reaction in vitro.

The objective of this investigation was to establish an in vitro counterpart to the in vivo graft-versus-host reaction. The generation in vitro of plaque-forming cells from spleen of rabbits previously immunized with sheep red blood cells could he completely suppressed by the simultaneous incubation of the memory cells with allogeneic circulating lymphocytes, sacculus rotundus, appendix or Peyer's patches cells (collectively referred to as SAPP effector cells). Spleen and lymph node cells were less effective in inhibiting the in vitro immune response whereas thymus and bone marrow exhibited no inhibitory activity. It was demonstrated that the inhibition of the generation of plaque-forming cells is not mediated by the release of cytotoxic antibodies directed to the memory spleen cells by the allogeneic lymphocytes. Following immunization with the appropriate antigen, allogeneic lymphocytes or SRBC so as to induce cell mediated and humoral responses respectively, the effector cells were shown to migrate to the thymus, whereas antibody forming cells were shown to migrate initially to the Peyer's patches. These results support the conclusion that the reaction represents an in vitro counterpart of the graft-versus-host reaction and imply a SAPP organ origin for the cells mediating this reaction in the rabbit