The most troublesome problem encountered during the sterile filtration of protein solutions is membrane fouling. This article presents our study on sterile filtration of a model protein, recombinant human growth hormone (rhGH). Scanning electron microscopy (SEMI analysis shows that 0.22-pm membranes, when used to filter the mannitol-formulated protein solution under a 0.35-bar transmembrane pressure, were plugged t o a great extent. When zinc ions were added t o induce aggregates, the fouling tendency of rhGH solutions increased with increasing amount and size of the aggregates, indicating that the aggregates present before filtration might be responsible for membrane fouling. However, repeated filtration of the same solution using a fresh filter each time cannot reduce membrane fouling, and all filtrates contain the same trace amount of hGH particulates as the prefiltered solution. Particulate size was determined to be between 0.03 and 0.15 p m by dynamic light scattering. Also, in view of the fact that protein formulations significantly affected the tendency of fouling with the same preexisting aggregates, it is likely that fouling was more attributed t o the aggregation taking place in the filter pores during filtration (secondary aggregation) than to the aggregates present before filtration. Adding a surfactant to or increasing the pH of the protein solution improves the filtration, whereas increasing ionic strength slows down the filtration. This result suggests that the balance of the protein's interaction and electrostatic repulsion plays an important role in the protein's fouling tendency. Many factors might change the microenvironment in the pores and disturb this balance. Those considerations and the aggregation tendency of rhGH in the filter pores will be discussed in detail separately. 0 1996