Immunoperoxidase Detection of Whey Protein Soils on Ultrafiltration Membranes ¹

An immunoperoxidase method was developed for detection of whey proteins in ultrafiltration (UF) membranes. A polyclonal antiserum was prepared against whey proteins in rabbits and coupled to horseradish peroxidase using sodium m-periodate as the oxidizing agent. The immunoperoxidase method was evaluated using a bench-scale, thin-channel UF unit equipped with polysulfone membranes for the concentration of whey to a 7.5 times concentration factor. Following rinse, membranes were washed for periods ranging from 0 to 60 min. The whey protein antibody-peroxidase conjugate was diluted in phosphate buffer and circulated in the system for 4 min. Unbound antibodies were removed by three consecutive washes with phosphate buffer containing 0.2% (v/v) Tween-20. Bound peroxidase conjugate was eluted by circulation of 2 M NaCl, pH 8.5, followed by distilled water, pH 7.4. To determine bound peroxidase, a chromogenic substrate was added to 2.5 ml portions of the eluants and absorbance recorded at 414 nm 30 min later. Nitrogen residues in the membranes varied from 0.061 to 0.317% and absorbance values in the respective eluents ranged from .381 to .582%. A significant correlation (r = 0.89, P<0.05) was found between nitrogen residue and bound peroxidase. The results suggested that the immunoperoxidase method has the desired sensitivity for non-destructive assessment of whey protein soils on bench-scale UF systems.