Study On IgY Extraction And Its Biological Activity

This paper mainly deals with the immunization to hen, the conditions of extraction, activity of IgY and the changes of specificity. It appears possible to use IgY for the fortification in functional food.Streptococcus mutans that caused dental caries in humans was used for immunization to hen. initial immunization was repeated once a week for four weeks, booster immunization was done 2 time from 10th week. After immunization, hen serum was collected and eggs were processed every week.A procedure was developed for extraction of IgY antibody from hen yolks by water dilution and freezing ethanol. The optimal recovery of protein was obtained when egg yolks were diluted 1:8 with water; precooled 95% ethanol was added to a final concentration of 60%; SephadexG-15 gel filtration or dialyzed against distilled water and Polyethylene glycol were used for condensation.Two extracting methods were compared. The procedure with ethanol precipitation was better , by this procedure, the protein samples included soluble protein(82.75%), water(17%) and ash(little), the content of soluble protein in total protein was 97% with yield of 12.6mg per milliliter egg yolk. A single yolk contains about 150mg IgY. After extracted by dextran sulphate, the content of protein was 19.02%, and the content of soluble protein was little.Antimicrobial assay showed that IgY resist the growing of E.coli and Bacillus Subtilis.the purity and molecular weight of IgY was determined by SDS-PAGE . reduced protein samples were analyzed by PAGE in a 10% acrylamide gel. After the protein was stained with a mixture of coomassie blue R-250 in 10% acetic acid-30% mcthanol, SDS-PAGE analysis yield two bands when the sample was reduced with 2-ME, IgY dissociated into a heavy chain and a light chain of 70KD and 40KD. After the protein was stained with Fuchsin basic, SDS-PAGR analysis yield one band.ELISA was applied to evaluate the specific activity of IgY. Specific activity has raised gradually from the 5th week and finally reached to the peak at the 8th week, serum and IgY remained high specificity against S.mutans after booster immunization at 10th week, specific activity can be obtained in a long time.