Preparation and characterization of antibody to galactosyl(alpha 1 leads to 4)galactosyl(beta 1 leads to 4)glucosylceramide.

Antiserum against galactosyl(alpha 1 leads to 4)galactosyl(beta 1 leads to 4)glucosylceramide (globotriaosylceramide, Gb3) was raised in rabbits by the administration of four weekly intramuscular injections of 1.5 mg of the purified glycoplipid along with bovine serum albumin and Freund's complete adjuvant. AntiGb3 activity was quantitated initially by immunoprescipitation employing Gb3 mixed with 100-fold excess of lecithin and cholesterol (1 : 1 or 1 : 2, by wt.) as antigen. Subsequently, complement fixation tests done with antigen preparations containing Gb3/lecithin/cholesterol (1 :6 :20, by wt.) showed antiGb3 titres of up to 1 : 8192. Fractionation of the antiserum by BioGel A5m chromatography indicated the antibody was an IgM immunoglobulin. The partially purified antibody stimulated complement-dependent release of glucose from glucose-containing liposomes prepared with shingomyelin/cholesterol/dicetylphosphate/Gb3 (molar ratio, 100 : 75 :11 :1). The antibody crossreacted with the trisaccharide, Gal(alpha1 leads to 4)Gal(beta1 leads to 4)Glc, but not with galactosylerceramide, lactosylceramide, GM1 ganglioside, globotetraosylceramide, digalactosyldiglyceride or a number of low molecular weight saccharides.