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|Title:||Preparation and characterization of immunogens for antibody production against metanephrine and normetanephrine|
|Authors:||Chan, E.C.Y. |
|Citation:||Chan, E.C.Y., Ho, P.C. (2002-08-01). Preparation and characterization of immunogens for antibody production against metanephrine and normetanephrine. Journal of Immunological Methods 266 (1-2) : 143-154. ScholarBank@NUS Repository. https://doi.org/10.1016/S0022-1759(02)00143-6|
|Abstract:||A two-step zero-length cross-linking procedure using active esters was successfully adopted for conjugating metanephrine (MN) and normetanephrine (NM) to bovine serum albumin (BSA). The protein was activated with water-soluble 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) in the presence of N-hydroxysulfosuccimide (sulfo-NHS), leading to the formation of active N-succinimidyl esters of some glutamic and aspartic acid carboxyls. The pertinence of this reaction for the coupling of these haptens to carboxylate groups was confirmed via reaction with a model compound, 2-hydroxybenzoic acid, and subsequent characterization using atmospheric pressure chemical ionization mass spectrometry (APCI-MS). Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) was used for the quantitative assessment of the hapten/protein ratios of these conjugates. This technique of conjugate characterization demonstrated greater resolution in molecular weight determination compared to nondenaturing polyacrylamide gel electrophoresis (native PAGE). Preliminary results from enzyme-linked immunosorbent assay (ELISA) and inhibition ELISA procedures using test antisera confirmed that the synthesized immunogens were highly antigenic and elicited specific antibody responses in BALB/c mice against the haptens. © 2002 Elsevier Science B.V. All rights reserved.|
|Source Title:||Journal of Immunological Methods|
|Appears in Collections:||Staff Publications|
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