Chemical ID of Visible Protein Agglomerates in Wet-Dispersion. AID

Oliver Valet, Ph. D., rapID, Berlin, Germany

Formation of protein agglomerates Development of protein based formulations exhibit a number of challenges as protein can undergo a variety degradation pathways. One of the most common changes that protein formulators experience is aggregation, resulting in formation sub-visible and visible particles that can negatively impact the drug’s performance. Protein agglomeration can be induced by various factors. Some common factors factors include temperature, light and shake stress; degradation of polysorbates followed by formation of fatty acids and a decrease in polysorbate concentration; silicone induced protein aggregation; and presence of tungsten particles in prefilled syringes.

Identification of protein agglomerates with the Single Particle Explorer Protein aggregation was induced by introducing an acidic tungstate solution to a protein solution. A small portion of the mixture containing a visible particle was then withdrawn with a pipette and transferred into a vial with particle free water. A picture of a particle in the vial was then taken with a photo camera (Figure 1). Inverted microscope was then used to capture a picture of the particle in situ (Figure 2). A portion of the solution (100 µL) containing the visible particle was withdrawn with a pipette and deposited into the wet-dispersion.AID. Sample was covered with a glass window and introduced to the Single Particle Explorer. Particle was manually located and photographed with 50x objective (Figure 2). Raman Spectroscopy was performed on the particle with 532 nm laser excitation, laser power of 100% and integration time of 30s. Particle spectrum matches reference spectrum of a protein with a rank value of 882.