Peptide-specific antibodies

MAbs produced by immunization with peptide may be screened first by ELISA to identify peptide specific antibodies. However, not all peptide specific antibodies will recognize the cell surface-expressed receptor. MAbs reactive with peptide must be subsequently screened by flow cytometry on receptor transfectants, cell lines, or leukocytes known to express the receptor in order to identify those which recognize native receptor. Alternatively, the fusion can be screened directly by flow cytometry to identify antibodies which recognize native receptor (see Subheading 3.3.2.),... 

Mintz KP, Mann KG. Detection of procollagen biosynthesis using peptide-specific antibodies. Matrix 1990 10 186-99. 

As noted, in avian gizzard myosin, all the HC204 and HC200 contain the 7-amino-acid insert. In avian aorta myosin, none of the heavy chains contains the insert. Whether some smooth muscles are composed of mixtures of inserted and noninserted isoforms, as suggested by the experiments of Babij (1993) and White et al. (1993), remains to be demonstrated conclusively through the use of peptide-specific antibodies. An alternative explanation for the apparent heterogeneity of some visceral tissues could be the presence of vascular myosin, which lacks the 7-amino-acid insert and which may originate from the blood vessels in the various viscera. 

Studies of PTM have been benefited from the recent advancements in mass spectrometry, the introduction of new software and Internet-based MS data search facilities, computer-assisted topology prediction for a variety of PTMs (visit http // ca.expasy.org/), chemical synthesis of modified peptides and proteins, development of modified peptide specific antibody, in vitro modification techniques, exploitation of other eukaryotic cells such as insect cells for protein expression [4, 5, 16, 32], and progress in affinity purification of modified proteins. 

Figure 7.3 The ER 1D5 peptide only binds to the ER 1D5 mAb (upper left and lower right). Other mAbs do not bind. The antibody abbreviations in the lower panel are Her2 11G5, and 9C2, human epidermal growth factor receptor type 2 clones 11G5 and 9C2 Mela-HMB45, melanocyte-specific antibody clone HMB45 Vimen V9, vimentin clone V9 Anti-LCA, anti-leukocyte common antigen clones PD7/26 and 2B11, combined as a cocktail Mouse poly IgG, mouse polyclonal IgG. Reproduced with permission from Sompuram et al.7...

Column A (Fig. 16.5) shows the baseline binding capability of each antibody for its peptide epitope. Prior experiments had established that each antibody only binds to its specific peptide and not the others.15 The presence of a colored spot on the glass slide indicates immunoreactivity. Each row contains a different peptide and antibody combination, as indicated by the legend to the left. The legend at the top of Figure 16.5 indicates that the peptides in column A are not treated in any way. 

Figure 19.3 The effectiveness of cBSA as an immunogen can be seen by the comparison of specific antibody response in mice to AV coupled to both nBSA cBSA. The quantity injected was standardized according to the amount of AV present. The cationized carrier results in higher concentrations of antibody produced against the peptide than the immunogen made with nBSA.

Posnett, D.N., McGrath, H., and Tam, J.P. (1988) A novel method for producing anti-peptide antibodies Production of site-specific antibodies to the T-cell antigen receptor b-chain. J. Biol. Chem. 263, 1719-1725. 

In another study, CPMV particles that displayed a peptide derived from the epidermal growth factor receptor variant III (EGFRvIII) elicited specific antibody responses in mice against the peptide and protected mice from tumor challenge [45]. 

Foot and mouth disease virus VP1 epitope Alfalfa leaf Mice developed specific antibody response to synthetic peptide,VP1 epitope, and intact FMDV particle. Immunogenic in mice when administered parenterally or orally. Mice protected against challenge with FMDV vims. 21, 23... 

Melnyk, O., Duburcq, X., Olivier, C., Urbes, R, Auriault, C., and Gras-Masse, H., Peptide arrays for highly sensitive and specific antibody-binding fluorescence assays. Bioconjugate Chem., 13, 713-720, 2002. 

An alternative way to validate the critical function eliciting the disease-relevant phenotype is the use of tool modulators these can be small molecules, peptides, or antibodies that may not have the properties to be considered a drug, but may display sufficient potency and selectivity to be used to interrogate the specific protein function in the relevant system. Such tools, however, are rarely available with the required characteristics to allow for a robust interpretation of the experiment. The exception is represented by the... 

Schmerr and Jenny established a CE-based immunoassay for the detection of prion protein (24). In this competitive assay, peptides derived from the prion protein and labeled with fluorescein were used. This allowed them to distinguish scrapie-infected brain preparations from noninfected. For identification, the ratio between the peaks resulting from the free and the com-plexed peptide with a specific antibody was used. The results were in agreement with other data on the brain preparations achieved by Western blot analysis. The CE-based assay provides the advantage of direct detection of the scrapie protein in blood and tissue preparations with high sensitivity. Furthermore, due to the small sample amount needed for analysis, the CE-based assay is applicable to the putative diagnostics of prion protein in body fluids. 

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