Why Does a Peptide Have to be Crosslinked to Another Protein?
Synthetic peptides used for antibody production are typically too small to have all of the necessary
information to induce the immune system to properly process them and in turn produce high titers
antibodies. Numerous techniques have been developed to utilize synthetic peptides but to
overcome the above limitations. Some companies utilize MAP (multiple antigen peptide) technology,
which allows the juxtaposition of 4 or 8 peptide molecules on a crosslinked lysine core. The MAP
technology, developed by Dr. James Tam, results in the production of antibodies to the peptides, but
in its simplest form, that used by companies offering custom antibody production, seems to have a
low incidence of antibodies produced that can recognize the native protein in situ. Dr Tam, in fact
has more recently developed a highly complex combination of lipid and multiple peptide copies that
seems to overcome these limitations. The costs of using the latter system are not justified given the
high degree of success using synthetic peptides conjugated to various immunogenic proteins. The
most commonly used of these carrier proteins is KLH, keyhole limpet hemocyanin, a very large and
very immunogenic protein that has been shown over many years to aid in the production of high titer
polyclonal antibodies. The primary sequence of KLH, with numerous exposed lysine residues,
allows for the covalent attachment of large numbers of peptide molecules. The resultant complex is
able to stimulate the immune system to produce antibodies against both the synthetic target peptide
as well as KLH itself. Thus it is also important that ELISA analysis be performed using synthetic
peptide conjugated to a different carrier protein, typically BSA. Finally, these is some evidence that
antibodies can also be produced that react to the crosslinked reagent used to couple the peptide to
KLH, and thus at 21st Century Biochemicals, we also utilize a different chemical crosslinker for the
attachment of peptide molecules to BSA than was used for coupling peptide to KLH. In this way,
ELISA can more accurately measure the presence of antibodies in serum specific for the target
peptide.
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