Abstract
| Original language | English |
|---|---|
| Pages (from-to) | 440-448 |
| Number of pages | 8 |
| Journal | Sensors and Actuators B: Chemical |
| Volume | 171-172 |
| DOIs | |
| Publication status | Published - 2012 |
| MoE publication type | A1 Journal article-refereed |
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Keywords
- Avidin
- biotin
- cysteine tagged
- non-specific binding
- self-assembled monolayer
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Cysteine-tagged chimeric avidin forms high binding capacity layers directly on gold. / Vikholm-Lundin, Inger (Corresponding Author); Auer, Sanna; Paakkunainen, M.; Määttä, J.A.E.; Munter, Tony; Leppiniemi, J.; Hytönen, V.P.; Tappura, Kirsi.
In: Sensors and Actuators B: Chemical, Vol. 171-172, 2012, p. 440-448.Research output: Contribution to journal › Article › Scientific › peer-review
TY - JOUR
T1 - Cysteine-tagged chimeric avidin forms high binding capacity layers directly on gold
AU - Vikholm-Lundin, Inger
AU - Auer, Sanna
AU - Paakkunainen, M.
AU - Määttä, J.A.E.
AU - Munter, Tony
AU - Leppiniemi, J.
AU - Hytönen, V.P.
AU - Tappura, Kirsi
PY - 2012
Y1 - 2012
N2 - 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.
AB - 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.
KW - Avidin
KW - biotin
KW - cysteine tagged
KW - non-specific binding
KW - self-assembled monolayer
U2 - 10.1016/j.snb.2012.05.008
DO - 10.1016/j.snb.2012.05.008
M3 - Article
VL - 171-172
SP - 440
EP - 448
JO - Sensors and Actuators B: Chemical
JF - Sensors and Actuators B: Chemical
SN - 0925-4005
ER -