Calculating the Mass of Subvisible Protein Particles with Improved Accuracy Using Microflow Imaging Data

Factors Governing the Accuracy of Subvisible Particle Counting Methods

“…[9][10][11] The most popular technologies available to date are NanoSight (nanoparticle tracking analysis [NTA]; NanoSight, Salisbury, UK) 12,13 ; Archimedes (resonant mass measurement [RMM]; Affinity Biosensors, Santa Barbara, CA) [14][15][16][17] ; coulter counter (CC, electrozone sensing) 18,19 ; and micro flow imaging (MFI) and FlowCam (FC, flow imaging microscopy; Fluid Imaging Technologies, Scarborough, ME). [20][21][22][23][24][25] However, each one of those emerging techniques measures different properties of the particulate matter to provide size and count information (e.g., NTA measures diffusivity, RMMedensity differences, CCeresistance changes, and FI captures images) Thus, it is important to note that each technique will be affected to a different extent by the same given properties of the sample and by specific artefacts. Even when those methodologies share a similar size range, the particle assessment performed by various methods will not necessarily compare.…”

“…In an attempt to evaluate the individual methods' performance a number of studies have examined different analytical aspects of the corresponding techniques, 12,13,[17][18][19][22][23][24][25][27][28][29][30][31][32][33] but to date, no systematic analysis which includes a comparison between all the different methods using a number of different standard systems, has been carried out. Recently, we reported on the factors governing the precision of subvisible particle measurement methods 26 in a detailed analysis of such an extensive analytical toolbox for subvisible particle characterization.…”

Reduced Subvisible Particle Formation in Lyophilized Intravenous Immunoglobulin Formulations Containing Polysorbate 20

“…We estimated the total mass of particles of size !2 mm based on a recently proposed ellipsoid volume method 25 (Fig. 4b).…”

“…A customized optical filter was used to remove air bubbles and the remaining particles were considered as protein particles. The total mass of protein in particles with size !2 mm was estimated using the ellipsoid-volume method described by Kalonia et al 25 …”

“…Subvisible particle characterization is particularly valuable in detecting trace amount of protein aggregates. [23][24][25] Particle counting methods provide a much more sensitive means by which to detect and quantify protein aggregation than traditional methods such as size exclusion chromatography (SEC). In 2 recent examples, particle formation was observed after monoclonal antibodies were subjected to freezethawing 23 or pumping, 24 but no loss of monomer or formation of soluble aggregates could be detected with SEC.…”