Do you need to know the amino-terminals of the heavy and light chains of your mAb?
We can sequence up to 30 residues from the N-termini of your antibody by N-terminal Edman degradation. Our experts perform N-terminal antibody sequencing, using two Procise 494 Edman sequencers.
You obtain the most accurate sequences on purified heavy chains and light chains. Typically, we reduce the antibody and separate the HC and LC by 1D SDS PAGE. The gel is then electroblotted onto a PVDF membrane. We now load the stained PVDF bands of the HC and LC onto to the Edman sequencer. Edman degradation is a cyclic procedure, which cleaves off one amino acid at a time. Afterwards it is identified by chromatography, and 10 cycles are sufficient to determine the N-terminus of most samples.
Antibodies are often modified in the N-terminals by partial formation of pyroglutamic acid. This blocks the terminal and thus prevents the Edman sequencing chemistry from working properly. We can remove the pyroglutamic acid by enzymatic treatment to enable the Edman sequencing.
Our N-terminal antibody sequencing service includes:
- First step is 1D SDS PAGE separation of the HC & LC.
- This is followed by blotting onto PVDF membrane.
- Next step is staining and cutting of the HC & LC bands.
- Then we perform Edman sequencing up to 30 amino acids.
- If needed, we can add de-blocking of pyroglutamic acid (optional).
- We deliver the sequencing report with easy to understand results, which you can use for your studies.
Download application note (pdf – 1 MB) Learn about the many applications of Edman degradation. – Then apply them to your studies for improved results.
Looking for specific residues? Then send us a request for a customized project.