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“…One part was stained with Coomassie brilliant blue (R-250) in a methanol-acid acetic mixture (31), whereas the other part was submitted to a transfer procedure adapted by Burnette (8) from Towbin et al (30). The electrophoretic transfer of proteins to nitrocellulose membranes (NCM; BA85 membrane filters, 0.45 ,um, Schleicher & Schuell, Dassel, Federal Republic of Germany) was done as previously described (23).…”
Section: Methodsmentioning
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“…One part was stained with Coomassie brilliant blue (R-250) in a methanol-acid acetic mixture (31), whereas the other part was submitted to a transfer procedure adapted by Burnette (8) from Towbin et al (30). The electrophoretic transfer of proteins to nitrocellulose membranes (NCM; BA85 membrane filters, 0.45 ,um, Schleicher & Schuell, Dassel, Federal Republic of Germany) was done as previously described (23).…”
Section: Methodsmentioning
“…HBsAg was obtained by extraction and purification from human sera [14] kindly provided by J. Pillot (Institut Pasteur, Paris, France). Briefly, plates (Nunc, Roskilde, Denmark) were coated overnight at 48C with HBsAg (subtypes ad and ay) at 10 g/ml.…”
Section: Detection Of Anti-hbs In Sera or Culture Supernatantsmentioning
“…The purification process of serum HBsAg involved a step of trypsin digestion in order to eliminate the tightly bound serum proteins [9]; such a purified antigen contained only traces of albumin and reacted with six different MoAbs specific to discontinuous a epitope of HBsAg [8], Proofs of anti-continuous specificity of the prepared MoAb allowing constant HBsAg detection were : (i) enhanced detection by EIA of reduced HBsAg compared with native antigen; (ii) total inhibition by reduced HBsAg and only partial inhibition by native HBsAg, Under the same conditions, reduced H BsAg was never detected by anti-discontinuous MoAb and reduced HBsAg did not inhibit anti-discontinuous MoAb [8]; (iii) reactivity with p24 polypeptide of the reduced HBsAg after SDS-PAGE, electrophoresis and immunoblotting.…”
Section: Discussionmentioning
“…Monoclonal antibodies (MoAb) recognizing a discontinuous epitope of the HBsAg group a specificity have previously been described [8], The anti-discontinuous MoAb IgGI chosen for this study recognizes highly purified HBsAg [9] in precipitation test, EIA and RIA, It did not recognize the reduced and alkylated purified HBsAG either in liquid phase or after adsorption on solid phase.…”
Section: Preparation Of Monoclonal Antibodies To Discontinuous Atid Cmentioning
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