Does anyone know which is the isoelectric point of an IgE? We are using an anti DNP-IgE and we think we are having some troubles because of the charge. Also, which buffer is better for this interaction (DNP-IgE)?
Thanks a lot
The heavy chain constant region of IgE (http://www.uniprot.org/uniprot/P01854) has a pI of just over 8 - obviously this does not include the variable region and light chain. Do you have a more complete peptide sequence?
Good luck with the question - please feel free to discuss further here.
Best wishes, Robin
In my opinion, the complete Peptide sequence of the IgE will say a lot about the pI. Now I am willing to learn also, is this sequence not source specific?
I guess you use the weird polyreactive clone SPE-7 IgE, or?
there are two partial sequnces of this clone and there are also several structural files:
https://www.ncbi.nlm.nih.gov/nuccore/KJ734990
https://www.ncbi.nlm.nih.gov/nuccore/KJ734989
so the complete fab fragment should have a pI of 8.4 (the heavy chain V region is very basic); lamba light chain should have 5.71 ( GenBank: AK008094.1 );
the heavy chain ( GenBank: LC031494.1 ) has 7.2;
so in total, the pI of the complete mab should be 6.5; But some carbs can change this.... depends on the source
the recombinant antibody is very unstable. only from hybridoma or selected eukaryotic expression systems, with full glycosilation it is somewhat stable. because the sequence is very weird which is quite typical for IgE. Not at all comparable to IgG.
I bet the clone purified from ascites will be really different in its biological activity. this is often the case for those close to germline, low affinity, polyreactive natural clones
- Badges
- Science method