National R. & D. Center for Egg Processing, Huazhong Agricultural University, Wuhan 430070, China.
An economical, effective and large-scale method for the preparation of immunoglobulin of chicken egg yolk (IgY) was developed. The method was based on an improved water dilution combining with polyethylene glycol 6000 (PEG6000) precipitation and anion-exchange chromatography. The egg yolk was diluted with 8 volumes of aseptic water, the pH of which was adjusted to 5.2 with 0.1 mol/L HCl, then the mixture was centrifuged at 5 000 x g after standing for 8 h at 4 degrees C. The crude IgY was obtained from the supernatant layer with the yield of 93.47%. The purification of IgY was achieved by 6% PEG6000 precipitation and DEAE-Toyo-pearl 650 M anion-exchange chromatographic packing. The optimum purified conditions by ion-exchange chromatography were as follows: equilibration with 0.05 mol/L phosphate buffer (pH 7.0) and elution with 0. 075 mol/L phosphate buffer (pH 7.0). The purity of the IgY fraction was consistently greater than 95% with high activity (73.77%) by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). This method is superior to the traditional ion-exchange chromatography that can't simultaneously obtain high purity and high recovery. Moreover, this method is suitable for a large-scale IgY preparation from chicken egg yolk.