A versatile dot-ELISA method with femtomole sensitivity for detecting small peptides.
Several protocols for conjugating peptides in situ to a protein carrier on paper, nitrocellulose, or nylon membranes were explored for their usefulness in dot-ELISA detection of the peptides. The most sensitive method in which peptide diluted in bovine serum albumin is applied to nitrocellulose, then fixed with glutaraldehyde, can detect several peptides, ranging from 4 to 38 amino acids in length, at the level of 2-10 fmol. Both immunohistochemical grade antisera and monoclonal antibodies have been used successfully. The method may be a useful alternative to radioimmunoassay since there is no requirement for radiolabelled peptide, or (for quantitation) for known quantities of unlabelled peptide. The method has been used to monitor, semiquantitatively, the fractionation of FMRFamide-like or CCK-like peptides from the nematode Ascaris, and to detect peptide-like immunoreactivities in tissue extracts.
Sithigorngul
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Department of Zoology, University of Wisconsin-Madison 53706.
Stretton
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Cowden
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D000818:Animals; D000911:Antibodies, Monoclonal; D001200:Ascaris / Q000032:analysis; D002766:Cholecystokinin / Q000032:analysis / Q000276:immunology; D002851:Chromatography, High Pressure Liquid; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D019835:FMRFamide; D007447:Invertebrate Hormones / Q000032:analysis; D009479:Neuropeptides / Q000032:analysis / Q000276:immunology; D010455:Peptides / Q000032:analysis*; D011817:Rabbits; D012680:Sensitivity and Specificity
D000911:Antibodies, Monoclonal; D007447:Invertebrate Hormones; D009479:Neuropeptides; D010455:Peptides; D019835:FMRFamide; D002766:Cholecystokinin