Correspondence work

 
  • Expression and purification of recombinant proteins (E. coli, yeast, insect cells, mammalian cells)
  • Biochemical analysis of biological samples (ELISA, Western blot, immunoprecipitation, etc.)
  • Enzyme activity measurement, stability measurement (Thermal shift assay, etc.)
  • Examination of protein crystallization conditions
  • Culture and mass expression of bacteria and fungi using jar fermenter
 

Protein expression purification test flow and action items

01
Hearing

ヒアリング

Highly specialized staff will hear about the problems that customers have. br / We will consider a solution.

  • Examination of expression / purification conditions on a small scale
  • Optimization and scale-up of purification conditions
  • Transfer of expression host

If you have such information, you will proceed promptly

  • Gene or amino acid sequence of the target protein
  • Information on the activity and stability of the target protein
  • Information on expression, purification, etc. performed in the past
  • Information on multimerization, decomposition, etc.
  • Presence or absence of homolog
02
Construction

Construction of expression vector

The expression host is determined and a vector expressing the target protein is constructed.

  • E. coli, yeast（～6L/回）
  • Insect cells（～6L/回）
  • Animal cells（Floating：～4.5L/回、fastening with glue：～3L/回）
Expression system owned by our company

  • Escherichia coli BL21 strain (including rare codon supplement strain)
  • Insect cell Sf-9 / baculovirus system
  • Animal cells CHO, HEK293
  • Yeast (Saccharomyces cerevisiae, methanol-utilizing yeast)

* We also support expression systems other than these. Please contact us individually.

03
Transfection/Transduction

Introduction of expression vector into cells

After culturing the cells and inducing the expression of the target protein, confirm the protein expression.

  • Expression confirmation (CBB staining, Western blot analysis)
  • Confirmation of solubility (CBB staining, Western blot analysis)
  • Activity confirmation (ELISA method, various enzyme activity measurement systems)
04
Selection

Selection of vector-introduced cells (establishment of cells)

The protein is purified according to the required degree of purification and the intended use of the protein.

  • Affinity chromatography (IMAC, GST, MBP, antibody, etc.)
  • Tag cleavage / removal with protease
  • Various chromatographys using FPLC system (gel filtration, ion exchange, etc.)
05
Analysis

Expression analysis of transgene in established cells

Using a part of the purified protein, check the concentration, activity, solution behavior, etc. as necessary.

  • Concentration test (ultraviolet absorption method, BCA method, Bradford method, etc.)
  • Activity confirmation (ELISA, various enzyme activity measurement systems)
  • Particle size distribution analysis by gel filtration chromatography

With Anewex R & D, you can do it!

  • I want to make a protein in E. coli that cannot be soluble and expressed in E. coli!
    (If you do not have a facility such as an animal cell expression system, but want to produce protein yourself)
  • We accept consideration of rewinding conditions from insoluble fractions.