Micro-Flow Imaging
MFI was used to detect particles up to 70 µm. A MFI5200 (ProteinSimple, Santa Clara, USA), equipped with a silane coated flow cell (1.41x1.76x0.1 mm) and controlled by the MFI View System Software version 2 was used. Prior to each measurement the system was flushed with purified water. The background was zeroed by using solvent and performing the optimize illumination procedure. Samples of 1 mg/mL adalimumab were analyzed without a predefined pre-run volume because of the limited amount. Flow rate was 0.17 ml/min and camera shot rate was 22 flashes per second. Data was analyzed with MFI View Analysis Suite version 1.2. For each product, stuck, edge, and slow-moving particles were removed by the software before analysis. Because no pre-run volume could be used, the data was recorded throughout the entire run but processed only in the time window from 0.7 to 1.7 min where, based on the trend chart option in the software, the measurement was stable for all samples. The equivalent circular diameter was calculated as described earlier (20).