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Enzyme-linked immunosorbent assay for riboflavin carrier protein and modified protein: application for epitope analysis.
Indian J Biochem Biophys ; 1992 Jun; 29(3): 277-81
Artigo em Inglês | IMSEAR | ID: sea-29006
ABSTRACT
Rabbit antibodies to native riboflavin carrier protein (RCP), are to a large extent directed towards the conformational epitopes and antibodies to disulphide bond reduced carboxymethylated riboflavin carrier protein (RCM-RCP) to the sequential epitopes. Taking advantage of this premise and in order to map the epitopes of RCP recognized by the antibodies, enzyme-linked immunosorbent assays were validated for RCP and RCM-RCP using the Avidin-Biotin system. The usefulness of these assays were illustrated when antigenicity of peptides derived from RCM-RCP following trypsinization were examined. Two major (T1,T2) and one minor peptide (T3) fractions were obtained when the tryptic peptides were fractionated on DEAE-cellulose. RCP has a blocked N-terminal. Tryptic peptides (T1 and T2) on microsequencing revealed the absence of an N-terminal amino acid, indicating that these fragments emanate from the N-terminal region of RCP. In support of this observation is the finding that antipeptide antibody to cRCP (10-24) of cRCP interacted with T1 as well as T2 indicating the presence of the sequential epitope (10-24) of cRCP in these fragments. In RCP-ELISA, only T2 displaced RCP and peptides T1 and T2 displaced RCM-RCP in RCM-RCP ELISA. Differences in the ability of these fragments (T1 and T2) to displace RCP and RCM-RCP reflect the subtle changes in the spatial structures of these epitopes in RCP and RCM-RCP.
Assuntos
Texto completo: DisponíveL Índice: IMSEAR (Sudeste Asiático) Assunto principal: Fragmentos de Peptídeos / Peptídeos / Proteínas de Membrana Transportadoras / Riboflavina / Ensaio de Imunoadsorção Enzimática / Dados de Sequência Molecular / Proteínas de Transporte / Sequência de Aminoácidos / Animais / Epitopos Idioma: Inglês Revista: Indian J Biochem Biophys Ano de publicação: 1992 Tipo de documento: Artigo

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Texto completo: DisponíveL Índice: IMSEAR (Sudeste Asiático) Assunto principal: Fragmentos de Peptídeos / Peptídeos / Proteínas de Membrana Transportadoras / Riboflavina / Ensaio de Imunoadsorção Enzimática / Dados de Sequência Molecular / Proteínas de Transporte / Sequência de Aminoácidos / Animais / Epitopos Idioma: Inglês Revista: Indian J Biochem Biophys Ano de publicação: 1992 Tipo de documento: Artigo