SAM #75N98026Q00506

Custom Antibody Development for N-homocysteinylated Lysine Detection in Biomedical Research

The National Institutes of Health Office of Logistics and Acquisition Operations (NIH OLAO), under the Department of Health and Human Services, is seeking specialized services for biomedical research: - Custom development and validation of a polyclonal antibody specific to N-homocysteinylated lysine (N-homocysteinylation), a post-translational protein modification - No commercial antibody currently exists for this modification, requiring a novel, custom solution - Scope of work includes: - Synthesis and purification of peptide antigens containing N-homocysteinylated lysine and matched control peptides - Conjugation of peptides to carrier proteins - Immunization of rabbits to generate polyclonal sera - ELISA screening and testing of sera on laboratory samples - Terminal bleed and purification of the working antibody - Validation of the antibody for use in characterizing proteins in mouse models of MMACHC deficiency (a rare genetic disorder) - The antibody will support research on rare genetic diseases and precision medicine - No specific OEMs or commercial vendors are named, as this is a custom antibody development requirement - Unique requirement: The antibody must be specific to N-homocysteinylated lysine, a modification for which no commercial detection reagent is available

Description

Requirements:

The proposed development of a novel post-translational modification (PTM)-specific antibody directly supports the U.S. Government's mission to advance biomedical research on rare genetic diseases, improve public health outcomes, and accelerate the development of precision medicine approaches

Currently there is no commercially available antibody against N-homocysteine-lysine (N-homocysteinylation) so we will need to have one generated. By generating it, we are able to provide specifications for the peptide sequence based on preliminary findings in our scientific models of disease insuring it will work effectively. Our objectives are as follows:  

Synthesize and purify peptide antigens with N-homocysteinylated lysine residues and matched control peptides with unmodified lysines.  Conjugate peptides to carrier proteins (e.g., KLH) for immunization  Immunize appropriate host species (rabbit for polyclonal antibody generation)  Generate polyclonal sera prior to immunization and 2 to 3 times after following antigen boost. Test sera on lab samples of N-homocysteinylated proteins from mouse models of MMACHC delicacy in our lab. Also perform ELISA screening of antibody on modified and unmodified peptide.   Terminal bleed on immunized animal and purification of working antibody from serum  With working antibody we will characterize the proteins affected by this aberrant posttranslational modification to better understand disease development and in future design targeted therapies for it. 

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