During formulation development of a therapeutic protein, combinations of buffers, pH and excipients need to be tested. As the protein bulk solution used for formulation development usually contains a buffer component at a defined pH and potentially one or more excipients already, this bulk requires to be processed. In case low concentrations of non-ionic surfactants, for example polysorbate 20, are already present in the bulk, the surfactant needs to be removed in lab-scale for further development use. The scope of the work was to study the behaviour of low concentrations of polysorbate 20 during membrane separation processes. The first part focuses on evaluating the behaviour of polysorbate 20 during a dialysis process, whereas the second part analyses concentration changes of polysorbate during a membrane concentration process using a stirred cell. The third part analyses potential membrane absorption of polysorbate at sterilizing-grade filters. In conclusion, it was found that polysorbate could not be significantly reduced during a dialysis process and accumulated during a membrane concentration process in unreproducable manner. During sterile filtration, no significant influence on the concentration of polysorbate was measurable. In any case, it is recommendable to quantify the concentration of polysorbate during critical membrane process steps in pharmaceutical industry.