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. 2008 Oct 15;24(2):334-7.
doi: 10.1016/j.bios.2008.05.003. Epub 2008 May 27.

Detection of human proteins using arrayed imaging reflectometry

Charles R Mace  1 , Christopher C Striemer, Benjamin L Miller
Affiliations

Detection of human proteins using arrayed imaging reflectometry

Charles R Mace et al. Biosens Bioelectron. .

Abstract

Assays built upon protein arrays are critical tools in determining the basic nature of biology, and have considerable promise in diagnosing human disease. These protein arrays aid in the elucidation of mapping pathway interactions, disease biomarker discovery, and regulatory processes. The solutions used in these experiments, including cellular lysate and serum, are inherently complex mixtures and are high in total protein content. Therefore, array-based assays must be robust and maintain a high level of selectivity and sensitivity. We report herein that arrayed imaging reflectometry (AIR), a label-free biosensing platform we have previously disclosed, is highly suitable for the detection of human proteins in complex solutions. In particular, we demonstrate array-based detection of cytokines in buffered solutions, and in undiluted human serum.

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Figures

Figure 1
Figure 1
Selectivity results for an array of five cytokine antibodies. In experiment A (light gray), the array was exposed to 1 µg/mL of TNFα and 1 µg/mL of IFNγ. In experiment B (dark gray), the array was exposed to the same TNFα/IFNγ mixture as in experiment A, but with an additional supplement of 500 ng/mL IL-5.
Figure 2
Figure 2
Detection of IFNγ. (a) Reflectance change profile (N=4) for target solutions of IFNγ in the range of 250 ng/mL to 25 pg/mL. (b) Image of the array used for detecting 250 ng/mL IFNγ. The top row corresponds to four anti-IFNγ spots. The bottom row, not readily visible in this image, lies directly beneath the top row and corresponds to four anti-human IgG control spots.
Figure 3
Figure 3
Detection of human proteins from undiluted serum. (a) Background, control image showing the array of anti-IFNγ (top left, mid bottom), anti-TNFα (bottom left, mid top), and anti-human IgG (right column). (b) Chip exposed to 100% serum. (c) Quantification of signal changes, corrected for background reflectance increases, for all probe spots (N=2)_on chips exposed to undiluted serum. The corrected IgG response represents saturation of the detector.
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