Cysteine-tagged chimeric avidin forms high binding capacity layers directly on gold

Inger Vikholm-Lundin, Sanna Auer, Maija Paakkunainen, Juha A Määttä, Tony Munter, Jenni Leppiniemi, Vesa P. Hytönen, Kirsi Tappura

Research output: Contribution to journalArticleScientificpeer-review

8 Citations (Scopus)

Abstract

Cysteine-tagged, genetically engineered avidin named ChiAvd-Cys and wild-type avidin form monolayers or bilayer structures when immobilised directly on gold. Non-specific binding can be reduced by a post-treatment of the avidin layers with a N-[tris(hydroxymethyl)methyl]-acrylamide (pTHMMAA) polymer. ChiAvd-Cys showed excellent activity when immobilised on gold. About 70% of the ChiAvd-Cys molecules were able to bind two biotinylated green fluorescent proteins (per avidin tetramer). Amino-biotinylated antibody F(ab′) 2 fragments could be bound to every 4th and 8th ChiAvd-Cys and wild-type avidin molecule, respectively, whereas on average one thiol-biotinylated antibody Fab′-fragment was bound to every ChiAvd-Cys. Antigen binding to the thiol-biotinylated Fab′-fragment bound to the ChiAvd-Cys/pTHMMAA layer was almost twice compared to that of the amino-biotinylated F(ab′) 2-fragments. The high antigen binding was due to a site-directed orientation of the thiol-biotinylated fragments. The ChiAvd-Cys/pTHMMAA layers offer high capacity that may be used to couple biotinylated compounds on biosensor surfaces.

Original languageEnglish
Pages (from-to)440-448
Number of pages9
JournalSensors and Actuators B: Chemical
Volume171-172
DOIs
Publication statusPublished - 2012
Publication typeA1 Journal article-refereed

Keywords

  • Avidin
  • Biotin
  • Cysteine tagged
  • Non-specific binding
  • Self-assembled monolayer

Publication forum classification

  • Publication forum level 2

Fingerprint

Dive into the research topics of 'Cysteine-tagged chimeric avidin forms high binding capacity layers directly on gold'. Together they form a unique fingerprint.

Cite this