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1. Cloning strategies

• Expression plasmid design, Species-specific codon optimization

2. Protein Expression in E. coli

• Single batch and fed batch, high cell density fermentation
• Inclusion body isolation and protein refolding
• Soluble protein purification in high yield

3. Purification of tagged Proteins

• One step purification with tags
• Remove tag with special enzymes

4. Further purification of protein

• Chromatography: Ion exchange, size exclusion, hydrophobic interaction, IMAC.
• Affinity: Dye chromatography, with immobilised proteins/peptides
• Refolding protein

5. Protein expression in mammalian cell lines

• Stable cell line production

6. Protein characterization

• Gel Electrophoresis (SDS-PAGE, Native, IEF, 2-D) and Coomassie blue or silver staining
• Protein concentration determination
• Western immunoblotting
• Absorption spectrum measurement
• N-terminal sequencing
• Peptide mapping
• Mass spectrometry
• Enzyme activity assay

We are able to use different expression systems like E. coli, yeasts (e.g. Pichia), or even mammalian cell lines for the expression of the proteins.

We are optimizing the expression conditions and therefore allowing the synthesis of proteins different scales – from reaction tubes and culture flasks to bioreactor. All Standard Cell Culture Equipment is available.

Isolation of cells

• Centrifugation, batch and flow-through
• Microfiltration

Cell disruption

• High pressure homogenizer
• Bead mills
• French press
• Sonifier

Clarifications and removal of cell debris, conditioning

• Ultracentrifugation
• Microfiltration
• Precipitation of proteins
• Aquous two phase systems

Protein purifications

• Chromatographic systems for the purification of proteins (ÄKTApurifier, ÄKTAexplorer, Smart-system)

Characterization

• SDS-PAGE, IEF, 2-D-electrophoresis, western blotting
• HPLC
• Edman-sequencing
• Maldi-Tof-MS/MS
• Electrospray with linear ion trap)