Adenovirus Particle Quantification in Cell Lysates Using Light Scattering

作者：Hohl Adrian; Ramms Anne Sophie; Dohmen Christian; Mantwill Klaus; Bielmeier Andrea; Kolk Andreas; Ruppert Andreas; Nawroth Roman; Holm Per Sonne^*

来源：Human Gene Therapy Methods, 2017, 28(5): 268-276.

DOI：10.1089/hgtb.2017.052

• adenovirus
• light scattering
• quantification
• downstream processing

Adenoviral vector production for therapeutic applications is a well-established routine process. However, current methods for measurement of adenovirus particle titers as a quality characteristic require highly purified virus preparations. While purified virus is typically obtained in the last step of downstream purification, rapid and reliable methods for adenovirus particle quantification in intermediate products and crude lysates to allow for optimization and validation of cell cultures and intermediate downstream processing steps are currently not at hand. Light scattering is an established process to measure virus particles' size, though due to cell impurities, adequate quantification of adenovirus particles in cell lysates by light scattering has been impossible until today. This report describes a new method using light scattering to measure virus concentration in nonpurified cell lysates. Here we report application of light scattering, a routine method to measure virus particle size, to virus quantification in enzymatically conditioned crude lysates. Samples are incubated with phospholipase A2 and benzonase and filtered through a 0.22 mu m filter cartridge prior to quantification by light scattering. Our results show that this treatment provides a precise method for fast and easy determination of total adenovirus particle numbers in cell lysates and is useful to monitor virus recovery throughout all downstream processing.

• 出版日期2017-10