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

Inger Vikholm-Lundin (Corresponding Author), Sanna Auer, M. Paakkunainen, J.A.E. Määttä, Tony Munter, J. Leppiniemi, V.P. Hytönen, Kirsi Tappura

Research output: Contribution to journalArticleScientificpeer-review

7 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 pages8
JournalSensors and Actuators B: Chemical
Volume171-172
DOIs
Publication statusPublished - 2012
MoE publication typeA1 Journal article-refereed

Fingerprint

Avidin
cysteine
Antigens
Antibodies
Gold
Cysteine
fragments
gold
Molecules
Sulfhydryl Compounds
thiols
Immunoglobulin Fab Fragments
Biosensors
antigens
Monolayers
antibodies
Proteins
Polymers
Acrylamide
Green Fluorescent Proteins

Keywords

  • Avidin
  • biotin
  • cysteine tagged
  • non-specific binding
  • self-assembled monolayer

Cite this

Vikholm-Lundin, I., Auer, S., Paakkunainen, M., Määttä, J. A. E., Munter, T., Leppiniemi, J., ... Tappura, K. (2012). Cysteine-tagged chimeric avidin forms high binding capacity layers directly on gold. Sensors and Actuators B: Chemical, 171-172, 440-448. https://doi.org/10.1016/j.snb.2012.05.008
Vikholm-Lundin, Inger ; Auer, Sanna ; Paakkunainen, M. ; Määttä, J.A.E. ; Munter, Tony ; Leppiniemi, J. ; Hytönen, V.P. ; Tappura, Kirsi. / Cysteine-tagged chimeric avidin forms high binding capacity layers directly on gold. In: Sensors and Actuators B: Chemical. 2012 ; Vol. 171-172. pp. 440-448.
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Vikholm-Lundin, I, Auer, S, Paakkunainen, M, Määttä, JAE, Munter, T, Leppiniemi, J, Hytönen, VP & Tappura, K 2012, 'Cysteine-tagged chimeric avidin forms high binding capacity layers directly on gold', Sensors and Actuators B: Chemical, vol. 171-172, pp. 440-448. https://doi.org/10.1016/j.snb.2012.05.008

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 journalArticleScientificpeer-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

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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 -

Vikholm-Lundin I, Auer S, Paakkunainen M, Määttä JAE, Munter T, Leppiniemi J et al. Cysteine-tagged chimeric avidin forms high binding capacity layers directly on gold. Sensors and Actuators B: Chemical. 2012;171-172:440-448. https://doi.org/10.1016/j.snb.2012.05.008