Method Optimization For Quantification Of Retroviral-Like Particles In Unprocessed Bulk Material By Thin-Sectioned Transmission Electron Microscopy

By Lara Strittmatter Ph.D., Sam Benigni, Gerard Marvin, Cole Rush, Stacie Fichthorn, Marian McKee Ph.D.

Transmission Electron Microscopy (TEM) is a crucial technique for the viral testing of unprocessed bulk (UPB) material, specifically for quantifying Retroviral-Like Particles (RVLPs), as required by ICH Q5A (R2). However, modern biomanufacturing has led to UPB with high protein content, which complicates sample preparation. The concentration of product protein alongside RVLPs results in a larger, highly compacted pellet that is difficult to properly homogenize, leading to poor accuracy and repeatability.

To overcome these challenges, a study explored modifying the centrifugation and sample volume steps. The results demonstrated that using a smaller starting sample volume (Half) combined with a long, low-speed centrifugation scheme significantly reduced the coefficient of variation (CV) for both RVLPs and gold nanoparticles (used as a homogenization tracker). This approach yielded a less compacted pellet, allowing for better homogenization and consequently, more accurate and precise particle quantitation. Learn more about the methodology and data, including updates to automated imaging and sampling integrity.