Immobilization of saccharides and peptides on 96-well microtiter plates coated with methyl vinyl ether-maleic anhydride copolymer

Ayano Satoh, Kyoko Kojima, Tamami Koyama, Haruko Ogawa, Isamu Matsumoto

Research output: Contribution to journalArticle

34 Citations (Scopus)

Abstract

We have previously reported a method to immobilize protein ligands on microtiter plates coated with methyl vinyl ether-maleic anhydride copolymer (MMAC) [Isosaki, K., et al. (1992) J. Chromatogr. 597, 123-128]. In this study, we improved the MMAC method to efficiently immobilize not only small ligands such as peptides and oligosaccharides, which could not be efficiently immobilized previously, but also heparin via its reducing end. Amino and hydrazino groups were introduced to MMAC-coated microtiter plate wells by coupling to acid anhydride groups of MMAC with 1,6-hexamethylenediamine and adipic acid dihydrazide, respectively. The amino groups introduced were allowed to react with peptides by use of divalent cross-linkers. Hydrazino groups were allowed to react with formyl groups of saccharides by reductive amination. Peptides and oligosaccharides were immobilized in a dose-dependent manner by these methods. In the case of the angiotensin peptide thus immobilized, the detection limit by monoclonal antibodies was as low as 0.1- 1 fmol peptide per well. Application of 20-200 nmol oligosaccharides to the well was sufficient to immobilize and subsequently detect lectins. Furthermore, heparin immobilized on the hydrazinocoated wells was successfully used for the binding assay of annexin IV.

Original languageEnglish
Pages (from-to)96-102
Number of pages7
JournalAnalytical Biochemistry
Volume260
Issue number1
DOIs
Publication statusPublished - Jun 15 1998

ASJC Scopus subject areas

  • Biophysics
  • Biochemistry
  • Molecular Biology
  • Cell Biology

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