High-Throughput Expression Screening in Mammalian Suspension Cells.

Proteins naturally expressed in eukaryotic organisms often require host chaperones, binding partners, and posttranslational modifications for correct folding. Ideally the heterologous expression system chosen should be as similar to the natural host as possible. For example, mammalian proteins should be expressed in mammalian expression systems.

Expression screening in mammalian suspension cells.

Proteins naturally expressed in eukaryotic organisms often require host chaperones, binding partners, and posttranslational modifications for correct folding. Ideally the heterologous expression system chosen should be as similar to the natural host as possible. For example, mammalian proteins should be expressed in mammalian expression systems.

Extended budded virus formation and induction of apoptosis by an AcMNPV FP-25/p35 double mutant in Trichoplusia ni cells.

An Autographa californica nucleopolyhedrovirus (AcMNPV) mutant (AcdefrT) isolated from virus-infected Trichoplusia ni (TN-368) cells produced plasma membrane blebbing and caspase-3-like activity late in infection. It also synthesized less polyhedra, but displayed enhanced budded virus formation in TN-368 cells. This phenotype resulted from dual mutations in p35 and FP-25.

Application of phage display to high throughput antibody generation and characterization.

We have created a high quality phage display library containing over 1010 human antibodies and describe its use in the generation of antibodies on an unprecedented scale. We have selected, screened and sequenced over 38,000 recombinant antibodies to 292 antigens, yielding over 7,200 unique clones. 4,400 antibodies were characterized by specificity testing and detailed sequence analysis and the data/clones are available online.

Multiplexed expression and screening for recombinant protein production in mammalian cells.

Background: A variety of approaches to understanding protein structure and function require production of recombinant protein. Mammalian based expression systems have advantages over bacterial systems for certain classes of protein but can be slower and more laborious. Thus the availability of a simple system for production and rapid screening of constructs or conditions for mammalian expression would be of great benefit.

Non-polar distribution of green fluorescent protein on the surface of Autographa californica nucleopolyhedrovirus using a heterologous membrane anchor.

Heterogeneous proteins can be displayed on the surface of the budded form of Autographa californica nucleopolyhedrovirus (AcMNPV) after fusion of the display protein to the AcMNPV major envelope glycoprotein, gp64. However, display is restricted to the poles of the virion and is relatively low level. To investigate the use of alternative membrane anchor sequences that would be compatible with virus surface display, we have constructed a display vector containing the gp64 signal peptide and a membrane anchor from the vesicular stomatitis virus (VSV) G glycoprotein.

Characterization of a truncated soluble form of the baculovirus (AcMNPV) major envelope protein Gp64.

A truncated tagged form of the Autographica californica multiple nuclear polyhedrosis virus major surface glycoprotein, gp64, has been expressed using the baculovirus expression system and purified to homogeneity by immune-affinity chromatography. The protein, which is responsible for virus-cell fusion, was a trimer in solution and retained this oligomeric form at pH 5, the pH of fusion. Circular dichroism spectroscopy indicated a protein with mixed alpha-helix and beta-sheet content that did not undergo significant change at pH 5.