Biochemical analysis of retroviral structural proteins to identify and quantify retrovirus expressed by an NS0 murine myeloma cell line

A subclone of the NS0 murine myeloma cell line, frequently used to produce recombinant monoclonal antibodies, was found by a transmission electron mic roscopy method to express a surprisingly high titer of 10(11) retroviral pa rticles per mi of culture supernatant. Infectivity assays showed a very low infectious titer with the restricted host range expected for a murine amph otropic retrovirus. A Western blot assay for the viral capsid protein was d eveloped to confirm the high titer values and provide a means for monitorin g batch consistency and virus removal during the purification process. Mass spectrometry of several of the viral Gag proteins demonstrated that the ce ll line appeared to produce at least two closely related retroviruses. N-te rminal sequencing of three of the Gag proteins demonstrated that these retr oviruses were members of the murine leukemia retroviral family. Western blo t detection with an antibody for the capsid protein gave a linear standard curve over the range of 0.1-3 ng per lane. This allows the detection of vir al titers as low as 6 x 10(7) virions per mi without the need to concentrat e the sample. The Western blot method has higher throughput and less variab ility than transmission electron microscopy methods and has potential for m onitoring viral titer and clearance during development of manufacturing pro cesses. (C) 2000 Elsevier Science B.V. All rights reserved.