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L. M. Woodward, S. Maswadi, R. D. Glickman, A. Oraevsky, A. Conjusteau, N. Barsalou, G. Zhong; Sensitivity of Optoacoustic Detection of Ocular Bacterial Antigen Using Targeted Gold Nanorods. Invest. Ophthalmol. Vis. Sci. 2008;49(13):4793.
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Recent investigations have demonstrated the capability of detecting ocular surface bacterial antigen utilizing a method based on laser optoacoustic spectroscopy and antibody-coupled gold nanorods (NR) as a contrast agent specifically targeted to the antigen of interest. (Woodward, et al. IOVS 48: E-Abstract 4247, 2007). We are investigating the sensitivity of this technique by determining the minimum concentration of detectable antigen.
Serial dilutions of C. trachomatis surface antigen (Ag) were prepared ranging from 1 mg/ml to 1 pg/ml. Using standard commercial reagents, a preparation of monoclonal antibody (Ab) specific to C. trachomatis surface Ag was conjugated to gold nanorods to be used as a targeted contrast agent with a strong optoacoustic signal (Oraevsky, et al. Nano lett; 7(7):1914-8, 2007). The dilutions of C. trachomatis surface Ag were adsorbed to the inside walls of detachable, flat-bottomed plastic micro-wells. A control sample containing albumin was used to determine specificity. An immunoaffinity reaction was then performed to allow binding of the targeted contrast agents. Optoacoustic signals were generated from the bound nanorods using methods previously described (Woodward et al, IOVS 48: E-Abstract 4247, 2007) by photothermal beam deflection at 520 nm (corresponding to one of the peak absorptions of the nanorods). The experiment was performed in a double-blind fashion.
An optoacoustic response was obtained that was proportional to the concentration of antigen contained in the micro-well. The assay’s limit of detection was approximately 1 pg/ml. No significant optoacoustic signal was generated from the micro-well containing albumin.
These findings indicate that antigen concentrations as low as 1 pg/ml can be detected in immunoaffinity reactions with the optoacoustic response from targeted contrast agents. Specificity of the targeted contrast agents was demonstrated as no optoacoustic signal was obtained from the albumin sample. These findings indicate that optoacoustic sensing of bacterial antigen with specific contrast enhancement is a sensitive and specific method for in vitro applications. Work is ongoing to compare this method to the current standards of ocular antigen detection.
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