Preparation of functionalized derivatives of inulin: conjugation of erythrocytes for hemagglutination and plaque-forming cell assays.

A method is described for preparing derivatives of alkali-stable polysaccharides for coupling to immunogen carriers or to sheep red blood cells (SRBC) for use in hemagglutination (HA) and plaque-forming cell (PFC) assays. Inulin, a beta (2 leads to 1)-linked polyfructosan was partially derivatized with carboxyl, aminoethyl or (p-aminophenyl)butyryl groups; the latter derivative was coupled to SRBC following diazotization. Optimal conditions for the sensitization of SRBC with inulin were given. The immunological reactivity of the inulin molecule was unaffected by the derivatization reactions, and high, reproducible anti-inulin HA titers for inulin-binding myeloma proteins were found using these specifically sensitized SRBC. The sensitized SRBC were stable for assays for over 2 weeks. Problems with spontaneous agglutination or distortion of sensitized SRBC, normally seen in other procedures, e.g., methods using stearoyl-inulin, were not encountered.