Enhancement of enzymatic colorimetric response by silver island films on high throughput screening microplates

• Published in: Journal of immunological methods 2014-09, Vol.411, p.43-49
• Main Authors: Abel, Biebele, Clement, Travis C., Aslan, Kadir
• Format: Article
• Language: English
• Subjects: Alkaline phosphatase; Alkaline Phosphatase - chemistry; Animals; bioassays; biosensors; Biotin-poly(ethylene-glycol)amine; Cattle; colorimetry; Colorimetry - methods; enzymatic reactions; enzymes; Enzymes, Immobilized - chemistry; High-throughput screening microplates; Horseradish peroxidase; Horseradish Peroxidase - chemistry; Membranes, Artificial; photography; Plasmonic nanoparticles; screening; Serum Albumin, Bovine - chemistry; Silver - chemistry; Silver island films; spectrophotometers; surface plasmon resonance; Surface Plasmon Resonance - methods
• ISSN: 0022-1759; 1872-7905
• DOI: 10.1016/j.jim.2014.06.005

In this study, we report the use of an enzyme-based hybrid platform, which is comprised of silver island films, enzymes (HRP and AP) and high-throughput screening (HTS) microplates, to enhance the colorimetric response of enzymatic reactions. The hybrid platform was designed in a two-step process: (i) deposition of SIFs onto HTS microplates with low, medium, and high loading (refers to the extent of the surface plasmon resonance peak of SIFs at 460nm) using Tollen's reaction scheme; and (ii) attachment of b-BSA or BEA as linkers for the immobilization of enzymes. The presence of SIFs within the wells of the HTS microplates was confirmed using an optical spectrophotometer and real-color photography. Control experiments, where SIFs were omitted from the surfaces were carried out to confirm the effect of SIFs on the enzymatic colorimetric response. Significant colorimetric signal enhancement was observed for HRP or AP on SIFs (high loading) deposited HTS microplates using b-BSA (up to ~3-fold for AP and ~6-fold HRP) or BEA (up to ~7-fold for both HRP and AP), as compared to our control samples. The observed increase in colorimetric response can be attributed to the nature of BEA, which exposes surface-bound enzymes to the substrate present in bulk more efficiently than b-BSA. This study proves that SIFs can serve as a valuable tool to improve the signal output of existing bioassays carried out in HTS microplates, which can be applicable to the field biosensors and plasmonics. SIFs can improve the signal output of existing bioassays carried out in HTS microplates, which can be applicable to the field biosensors and plasmonics. [Display omitted] •High throughput screening microplates are modified with silver island films.•Biotinylated polymer and a protein were used as linkers.•Colorimetric response for HRP can be increased 7-fold.•Colorimetric response of AP can be increased 6-fold.