Moxtek has developed cost effective and reliable methods to obtain uniform wafer scale metallic nanohole array patterned biochips. Here their use as fluorescence enhancing substrates for Cy3 based protein microarrays is demonstrated. The structures, utilizing highly focused and enhanced optical fields, greatly boost the fluorescence SNR, allowing for the detection of antigen concentrations 3+ orders of magnitude lower than current commercial standards.

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Standard microarray based fluorescence protocols were carried out on an Al nanohole array (NHA) patterned slide obtained from a fully patterned 200mm diameter glass wafer. The process involved spotting with 4 duplicates of the three antigens Human IgG, IgM, and biotinylated BSA at concentrations of 0.1mg/ml – 0.1ng/ml over x10 serial dilutions. Additional positive and negative control spots were also added and the process was repeated over 16 subarrays on a single slide. The subarrays were then assayed with the corresponding Cy3 labeled antibodies of Human IgG, IgM, and Streptavidin at a concentration of 0.1ug/ml. Fluorescence results were then averaged from multiple spots over multiple subarrays.

This process was also carried out on two commercial standards: Schott Nexterion E and HiSens E slides, which served as baseline for comparison. The resulting fluorescence images and quantitative results showed that the Moxtek Al NHA slide offered a very clear advantage, especially as the antigen concentration was decreased. In fact, the Moxtek slide offered at least 11x and 25x enhancements of the fluorescence SNR over the Schott HiSens E and Nexterion E slides respectively at these lower concentrations. Aluminum nanohole array substrates are now available. Please contact Moxtek for additional details.