Sensitive fluorescence-based method for the rapid determination of polysorbate-80 content in therapeutic monoclonal antibody products.

ABSTRACT

A sensitive and effective method has been developed for the rapid determination of polysorbate-80 content in therapeutic monoclonal antibody (mAb) products. The method is based on the detection of the fluorescence emission of 4,4'-dianilino-1,1'-binaphthyl-5,5'-disulfonic acid dipotassium salt (bis-ANS) enhanced by the presence of polysorbate-80. The developed method includes two approaches. One requires removal of the mAb from solution prior to analysis, while the other requires only simple sample dilution. The limits of detection and quantitation, calculated from the calibration curve generated in the absence of mAb-A, were 1.5 and 4.7 parts per million, respectively. Given the comparable linear range and linearity of the linear line between the solutions, with or without mAb, the limit of detection and quantitation is assumed to be similar. The dilution method is not only fast and simple in terms of sample preparation, but it is also particularly useful for analyzing the level of polysorbate-80 contained in highly concentrated mAb products. However, given that this method does require availability of polysorbate-80-free materials of mAb for preparation of calibration standards, the protein removal method may be useful for the cases where appropriate protein materials for standard preparation are limited or unavailable.