Medical Biotechnology Research & Development, Research Council-Tamilnad Hospital Academic Trust, Chennai, India
Russ. J. Immunol. 1999, July 15, 4 (2), 131-140.
Chicken egg yolk is an abundant, yet inexpensive source of polyclonal antibodies. However, removal of the contaminating lipids and lipoproteins from immunoglobulins of egg yolk (IgY) is considered to be a laborious task during the purification. Liposomal asialoGM1 was used as a model immunogen for raising polyclonal IgY and tested with various methodological approaches. To overcome certain difficulties posed during IgY purification, several detergents (cationic, anionic and non-ionic) were tested for their effect on yolk protein and lipoprotein separation during water dilution processing of IgY. Cetrimide showed a pronounced effect on improving the immunoglobulin separation with only less than 3% of the lipid contaminants as compared to other detergents. The least effective detergent was sodium dodecyl sulphate with 14-35% contaminating lipids. The usefulness of egg yolk acetone powder for long-term storage prior to IgY purification was also determined. The use of acetone precipitated proteins for IgY processing showed a 25-30% loss in antigen specific immunoreactivity despite of an optimum total immunoglobulin recovery of 80%. A combination of delipidation of yolk with 50% acetone and treatment of the resultant acetone precipitated protein with a low ionic buffer (pH 5.5) containing 200 mcM of cetrimide yields electrophoretically homogeneous IgY with an optimum total immunoglobulin recovery of 85-90%. The antigen specific immunoreactivity of anti-asialoGM1 IgY was preserved well during the cetrimide treatment and was found to be higher than all other recovery processes, as tested by radial immunodiffusion and immunoblot assays.