Production and purification of Fab' fragments from chicken egg yolk immunoglobulin Y (IgY)

Methods were described for the production of Fab and Fab' fragments from chicken egg yolk IgY also referred to as IgG by papain and pepsin digestion respectively. Pepsin digestion was found to be suitable for the large scale preparation and purification of Fab'. Optimum yield of Fab' was obtained after peptic digestion of IgY at pH 4.2 for 9 h at low NaCl concentration. This condition led to the complete digestion of pFc' fragment leaving only the Fab' fragment. By combination of ultrafiltration and anion exchange, and conditions which allowed binding of the small amount of contaminants in the digest to the anion exchange column, pure Fab' fragments were easily obtained in the eluent. The advantage of this approach is that a small column could be used to purify large amount of protein, therefore, improving the efficiency of purification. The Fab and Fab' fragments appeared to be similar on the basis of their molecular weights as determined by SDS-PAGE, reaction of identity in immunodiffusion assay and similar antigen binding activities as shown by ELISA.