Recombinant Protein Production

Recombinant Protein Production refers to a series of steps, starting with an amino acid sequence and resulting in a concentrated sample of proteins with that sequence. The process involves Recombinant Protein Expression, separation, Protein Purification and concentration. In the case of recombinant protein purification or recombinant antibody purification, the starting material is generated by an expression system (cells that have been engineered using recombinant DNA technology to produce the desired protein abundantly).

Protein Sequencing prior to Recombinant Protein Production

De novo protein sequencing is an important step to confirm the sequence or identity of the protein or antibody of interest before, during or after a protein production and purification process. There are several applications, including:

  1. Prior to Recombinant Protein Expression, the sequence of the desired protein or antibody must be known with 100% certainty. If the objective is to produce more of a commercially interesting protein sample, but the identity or sequence of the protein/antibody is not completely certain, de novo protein sequencing can determine the sequence without reference to any DNA sequence.
  2. When designing an Antibody or Protein Production workflow, de novo protein sequencing can be used to confirm the effectiveness of the Protein Purification process. The antibody or protein sequence and characteristics can be determined directly from the purified Antibody sample at any stage of the workflow prior to investing in scaling up production.
  3. In research applications, Protein Sequencing can be used after protein purification is completed to understand and characterise Proteins or Antibodies of interest
  4. In any application, Antibody Sequencing can be used to understand the identity and sequence of a sample of antibody in order to understand how to produce more.

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Recombinant Protein Expression

See Protein Expression.

Recombinant Protein Purification

There are two principal types of protein purification: analytical and preparative. Preparative purification produces large volumes of proteins or antibodies and is an important part of protein production. However, recombinant protein production can also refer to protein expression.

Depending on the degree of purification required, one can make use of several properties of the protein to separate it from others:

  • Protein Hydrophobicity – using Hydrophobic Interaction Chromatography (HIC)
  • Charge – Ion Exchange Chromatography
  • Size – filtration, gel electrophoresis, SDS Page (which also involves charge)
  • Binding affinity – depending on the protein of interest, this could involve lectin affinity, ligands, metal affinity, antibody-antigen affinity
  • HPLC is a common technique

These steps may also be used sequentially.

Downstream processes often involve filtration to increase the concentration of the protein of interest.

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