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Method Article
Sandeep Kumar Vashist
HSG-IMIT - Institut für Mikro- und Informationstechnik; Laboratory for MEMS Applications, Department of Microsystems Engineering -IMTEK, University of Freiburg, Georges-Koehler-Allee 103, 79110 Freiburg, Germany
Corresponding Author
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A rapid and highly-sensitive procedure has been developed for surface plasmon resonance (SPR)-based immunoassay (IA) for human fetuin A (HFA). It employs a highly-simplified antibody (Ab) immobilization strategy, which only involves sequentially the dispensing of anti-HFA capture Ab diluted in 1% (v/v) 3-aminopropyltriethoxysilane (APTES) onto a potassium hydroxide (KOH)-treated gold (Au)-coated SPR chip followed by its incubation for 30 min. The devised Ab immobilization strategy enables the leach-proof binding of capture Ab, resulting in highly reproducible and cost-effective HFA IAs. The SPR IA detected HFA with a dynamic range, limit of detection, and analytical sensitivity of 0.3-20 ng mL-1, 0.7 ng mL-1 and 1 ng mL-1, respectively. The detecting chip can be easily regenerated after each analyis by treatment with 10 mM glycine-HCl, pH 2.0. The Ab-bound SPR chip can be stored at 4°C for up to 4 months without any significant loss of its functional activity. The developed IA procedure has been employed to detect HFA spiked in diluted human whole blood and plasma. Moreover, the developed SPR IA has high analytical precision as its results are correlated well with those of conventional HFA sandwich ELISA.
Keywords
Surface plasmon resonance, immunoassay, one-step antibody immobilization, 3-aminopropyltriethoxysilane, human fetuin A
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The authors declare no competing financial interests.
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Associated Publications
Surface plasmon resonance-based immunoassay for human fetuin A
by S. K. Vashist, E. M. Schneider, and J. H. T. Luong
The Analyst (29 April, 2014)
An open repository of community-contributed protocols sponsored by Nature Research.
Learn more about Protocol Exchange
Method Article
Sandeep Kumar Vashist
HSG-IMIT - Institut für Mikro- und Informationstechnik; Laboratory for MEMS Applications, Department of Microsystems Engineering -IMTEK, University of Freiburg, Georges-Koehler-Allee 103, 79110 Freiburg, Germany
Corresponding Author
Version History
CURRENT STATUS: Posted
Version 1
Posted 30 Apr, 2014
No community comments so far
Metrics
Comments: 0
HTML Views: ...
License:
This work is licensed under a CC BY-NC 3.0 License. Read Full License
A rapid and highly-sensitive procedure has been developed for surface plasmon resonance (SPR)-based immunoassay (IA) for human fetuin A (HFA). It employs a highly-simplified antibody (Ab) immobilization strategy, which only involves sequentially the dispensing of anti-HFA capture Ab diluted in 1% (v/v) 3-aminopropyltriethoxysilane (APTES) onto a potassium hydroxide (KOH)-treated gold (Au)-coated SPR chip followed by its incubation for 30 min. The devised Ab immobilization strategy enables the leach-proof binding of capture Ab, resulting in highly reproducible and cost-effective HFA IAs. The SPR IA detected HFA with a dynamic range, limit of detection, and analytical sensitivity of 0.3-20 ng mL-1, 0.7 ng mL-1 and 1 ng mL-1, respectively. The detecting chip can be easily regenerated after each analyis by treatment with 10 mM glycine-HCl, pH 2.0. The Ab-bound SPR chip can be stored at 4°C for up to 4 months without any significant loss of its functional activity. The developed IA procedure has been employed to detect HFA spiked in diluted human whole blood and plasma. Moreover, the developed SPR IA has high analytical precision as its results are correlated well with those of conventional HFA sandwich ELISA.
Figure 1
Figure 2
Figure 3
Figure 4
Figure 5
The authors declare no competing financial interests.
Loading...
Associated Publications
Surface plasmon resonance-based immunoassay for human fetuin A
by S. K. Vashist, E. M. Schneider, and J. H. T. Luong
The Analyst (29 April, 2014)