Effective and efficient characterization of Chinese hamster ovary production cell lines using automated intracellular staining and statistical modeling.

Mammalian cell line development is critical to bioproduct manufacturing. Success requires selecting a line with desirable performance characteristics, including consistent expression throughout the proposed manufacturing window. Given the genetic and phenotypic flux inherent to immortalized lines such as Chinese hamster ovary cells, clonally-derived cell line characterization is vital.

Improving the efficiency of CHO cell line generation using glutamine synthetase gene knockout cells.

Although Chinese hamster ovary (CHO) cells, with their unique characteristics, have become a major workhorse for the manufacture of therapeutic recombinant proteins, one of the major challenges in CHO cell line generation (CLG) is how to efficiently identify those rare, high-producing clones among a large population of low- and non-productive clones. It is not unusual that several hundred individual clones need to be screened for the identification of a commercial clonal cell line with acceptable productivity and growth profile making the cell line appropriate for commercial application. This inefficiency makes the process of CLG both time consuming and laborious.

5'AMP-activated protein kinase alpha deficiency enhances stress-induced apoptosis in BHK and PC12 cells.

5'AMP-activated protein kinase (AMPK) activation occurs under a variety of stress conditions but the role of this enzyme in the promotion or inhibition of stress-induced cell death is unclear. To address this issue, we transformed two different cell lines with shRNA-expressing plasmids, targeting the alpha subunit of AMPK, and verified AMPKalpha downregulation. The cell lines were then stressed by exposure to medium without glucose (PC12 cells) or with the viral thymidine kinase-specific DNA replication inhibitors: acyclovir, penciclovir and ganciclovir (herpes simplex virus thymidine kinase-expressing Baby Hamster Kidney cells).

Sample preparation for two-dimensional gel electrophoresis.

The choice of sample preparation protocol is a critical influential factor for isoelectric focusing which in turn affects the two-dimensional gel result in terms of quality and protein species distribution. The optimal protocol varies depending on the nature of the sample for analysis and the properties of the constituent protein species (hydrophobicity, tendency to form aggregates, copy number) intended for resolution. This review explains the standard sample buffer constituents and illustrates a series of protocols for processing diverse samples for two-dimensional gel electrophoresis, including hydrophobic membrane proteins.

Temporal pattern of herpes simplex virus type 1 infection and cell death in the mouse brain stem: influence of guanosine nucleoside analogues.

Levels of bystander death occurring in herpes simplex virus type 1 (HSV-1)-infected mouse brain stems were studied, as well as the extent to which bystander death is influenced by guanosine nucleoside analogue treatment. Consecutive sections from brain stems of HSV-1-infected mice were stained alternately for (i) viral infection and (ii) cell death (TUNEL assay). Virus antigen was detectable in brain stems on day 3 of infection, while TUNEL staining was comparatively lower.