Protein Services

All West Bioscience customized protein expression and purification are carried out in our California facility to insure highest quality and best customer satisfaction. We have been a leader in providing high quality recombinant protein expression and purification services, especially in customized protein design area. We delivered thousands proteins to our customers worldwide.

Pros and Cons of E. coli system

Pros

Potentially very high expression levels
Low cost
Rapid growth
Scalable
Simple transformation protocols
Many parameters can be altered to optimize expression

Cons

Inefficient disulfide bond formation
Poor folding of proteins in the cytoplasm
Inclusion body formation
In vitro refolding protocols inefficient – may negate advantages
Codon usage different to eukaryotes
Minimal post-translational modifications
Endotoxin

Cloning

WE will clone your insert into an expression vector by either a standard subcloning or a PCR cloning. will be used to transfer the gene of interest into the expression vector. Clones will be fully sequenced for verification.

Screening of Expression

The expression vector containing the gene of interest will be transformed into an expression strain of E. coli for protein production. Several individual clones will be screened in a small-scale induction experiment. The expression results will be analyzed by SDS/PAGE and Western blot.

Optimization of Expression

The expression condition will be optimized by time of induction, temperature and media condition. The results will be confirmed by SDS-PAGE and Western blot.

Protein Characterization

A small-scale culture will be tested and the expressed protein will be characterized for cellular location, solubility, yield and accessibility of purification tags. A detailed strategy for producing and purifying the protein will be designed.

Protein Production

Large-scale production in liters volume will be made using shake flasks. If you prefer purifying protein by your own, you will receive the cell pellets and a complete report with the SDS-PAGE and Western blot analysis results.

Protein Purification

Several methods can be used to purify your protein such as inclusion body preparation or affinity column purification.

Pros and Cons of insect system

Pros

Good expression levels (especially for intracellular proteins)
Relatively rapid growth
Efficient protein folding
Moderately scalable
Extensive post-translational modification of proteins
Glycosylation more like mammalian cells
Endotoxin-free

Cons

Expensive culture media
Large volumes of virus needed on scale-up, time consuming
Inefficient processing of pro-peptides in secretory pathway
Glycosylation still different to mammalian cells
Viral infection leads to cell lysis and potential degradation of expressed proteins

Cloning

West Bioscience will clone your insert into a baculovirus transfer vector via subcloning or a PCR cloning. Clones will be fully sequenced for verification.

Recombinant Virus Production

West Bioscience will cotransfect a baculovirus transfer vector into Sf9 cells using linearized DNA from Autographa californica nuclear polyhedrosis virus (AcMNPV). Virus titer will be amplified 2-3 times to generate high-titer virus stocks.

Optimization of Expression

Recombinant protein expression will be optimized with a time-course study of infection, varying the multiplicity of infection (MOI). At each time point, the expressed protein will be analyzed by SDS-PAGE and/or Western blot. The optimal conditions for protein production will be determined.

Protein Characterization

A strategy for producing and purifying the protein will be decided on the protein characterization. At this step protein from small-scale culture will be characterized for cellular location, solubility, yield and accessibility of purification tags.

Protein Production

Production of the recombinant protein will be scaled up to liter volumes. If you prefer purifying the protein by your own, you will receive the cell pellets or conditioned media and a complete project report with the SDS-PAGE and/or Western blot analysis.

Protein Purification

West Bioscience will purify your recombinant proteins using an affinity column. Large-scale purification is also available.

Pros and Cons of mammalian system

Pros

Good expression levels
Moderately scalable
Suspension-adapted cells facilitate scale-up
Efficient protein folding
Good for secreted proteins
All post-translational modifications
Endotoxin-free
Rapid route to protein (transient)

Cons

Expensive culture media
Complex growth requirements
Transfection reagents can be expensive at scale (transient)
Large amounts of plasmid DNA for scale-up (transient)

Cloning

West Bioscience will clone your insert into the desired mammalian expression vector using standard subcloning or PCR cloning. Clones will be sequenced for verification.

Optimization of Expression

293 cells grown in serum free medium will be transiently transfected with the mammalian expression vector. Small-scale protein production will be optimized by time course of expression. The results will be analyzed by SDS-PAGE and Western blot. The yield and best expression condition can be determined and an expression and purification strategy can be designed.

Protein Production

Large-scale production in liters can be made. If you prefer, cell pellets or conditioned media can be delivered. A complete report with the SDS-PAGE and Western blot analysis will be made.

Protein purification

We will purify your recombinant proteins using an affinity column.

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