igBLAST and Antibody Protein Sequencing

What is igBLAST?

IgBLAST is a sequence analysis tool for antibody variable domain sequences. It matches matches to the germline V, D and J genes, details at rearrangement junctions, the delineation of IG V domain framework regions and complementarity determining regions (CDR). Understanding the sequence of your antibody is an important, and often critical step in antibody research. igBLAST provides a useful way to compare your antibody sequence to reference sequences.

Options for Antibody Sequencing and igBLAST

To determine the antibody’s sequence, you have two options:

  1. DNA sequencing techniques can be used, if you have access to the cell-line/hybridoma, to determine the antibody sequence
  2. You can use antibody sequencing (a.k.a. Antibody Protein Sequencing) techniques to sequence your antibody if you just have an antibody sample or you wish to understand post-translational modifications, phosphorylation, glycosylation, etc.

In both cases, igBLAST can be used in post-analysis.

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Procedure for Antibody Protein Sequencing

Antibody Protein Sequencing is generally achieved in a bottom-up manner by the following steps: (1) Enzymatically digest the protein into shorter peptides. (2) Conduct mass spectrometry measurement to produce the tandem mass spectrometry (MS/MS) data for the peptides. (3) Derive the sequence of each shorter peptide from the MS/MS data (a.k.a. De novo peptide sequencing). (4) Assemble the peptides sequences back to the long protein sequence.

igBLAST and de novo antibody protein sequencing

Antibody Protein Sequencing (a.k.a. Antibody de novo sequencing) methods can determine complete antibody sequences with high confidence and without reference to a DNA reference sequence. Read more about how Antibody Protein Sequencing works. Once an antibody has been sequenced in this way, its sequence can be used with igBLAST to find homologous antibodies.

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