Here is the result of my antibody survey. I had asked people for whether
they had good/bad experience with antibodies raised against fusion proteins
or peptides.
Fusion worked: 13 cases
Fusion didn't work: 4 cases
6xHIS worked: 2 cases
6xHIS didn't work: 1 case
Peptide worked: 7 cases
Peptide didn't work: 3 cases
I counted only those cases where it was clear whether the antibody works or
doens't work on whole-mounts and excluded cases that were "doomed" from the
beginning (i.e. insoluble peptides, for example).
In summary the success rate for anti fusion AB (75%) is about the same as
for antipeptide antibodies (70%).
It seems to be a critical parameter how to choose the peptide: Best if from
N- or C-terminus (= mostly exposed parts of the protein) + soluble as
judged by eye , i.e. polar amino acids preferable (= high probablity that
on surface of native protein + best to couple to carrier).
In regard to my epitope survey I got very few responses. It seems that both
HA and myc epitope tags work fine.
Oliver
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Oliver Hobert
Harvard Medical School/Department of Genetics
Mass. General Hospital/Department of Mol. Biol.
Ruvkun-Lab
Wellman 8
Boston, MA 02114
Fax: 1-617-726 6893
Tel: 1-617-726 5983
e-mail: hobert at molbio.mgh.harvard.edu
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