Purpose: Flow-imaging microscopy (FIM) is used to visualize and quantify micrometer-sized particles in biopharmaceutical during various stages of drug product development. Traditional FIM systems resolve particles in size down to ca. 1 µm by using a planar flow-through cell in front of a light source that is inspected by a magnified digital camera.
Recently, there is the possibility to extend the size range of FIM towards the submicron-size range. The use of immersion oil and higher magnification allows a higher resolution, which enables the detection of particles in the hundredths of nanometer-size range. In addition, more detailed images of small particles that are captured in only few pixels with traditional FIM systems are collected so that it is possible to visualize even submicron particles.
We highlight Coriolis’ first insights into capabilities and limitations of the new technique for the characterization of pharmaceutically relevant samples, which contained both amorphous, proteinaceous particles and silicone oil droplets.
Methods: Three different biopharmaceutical drug products (DP1: IgG lyophilized in vials, DP2: IgG in pre-filled syringes (PFS), and DP3: small protein in PFS) were analyzed on the recently released FlowCam Nano (Fluid Imaging Technologies) system, using 40x magnification, immersion oil optics, and 50-µm flow cell. The results were compared to traditional systems including MFI 5200 (Protein Simple) and FlowCam 8100 ALH (Fluid Imaging Technologies) with 4.88x (100-µm SP3 flow cell) and 10x magnification (80-µm FOV flow cell), respectively.
Results: The use of immersion oil and higher magnification allowed a higher resolution, which enabled the detection of particles in the size range of ca. 0.3 mm and larger. In addition, more detailed images of small particles that are captured in only few pixels with traditional FIM systems were collected, which allowed visualization of particles < 5 µm and submicron particles.
Finally, because of the small analyzed volume, FlowCam Nano would require a larger sample volume to count statistically significant numbers of micron particles.
Conclusion: In this first feasibility study, without yet fully optimized measurement conditions for sizing and counting, promising results of relevant biopharmaceutical samples containing both amorphous protein aggregates as well as silicone oil droplets were obtained, which extent the measurement range of FIM into the submicron-size range.
Potentially, the technique supports the evaluation of the pharmaceutical relevance of submicron particles for the safety and efficacy of biopharmaceutical drug products.