TUTCRIS - Tampereen teknillinen yliopisto

TUTCRIS

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

Tutkimustuotosvertaisarvioitu

Yksityiskohdat

AlkuperäiskieliEnglanti
Sivut440-448
Sivumäärä9
JulkaisuSensors and Actuators B: Chemical
Vuosikerta171-172
DOI - pysyväislinkit
TilaJulkaistu - elokuuta 2012
OKM-julkaisutyyppiA1 Alkuperäisartikkeli

Tiivistelmä

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.