Nimble Screen

For the routine screening of larger probe sets, an intermediate level of multiplex sample hybridization has been developed, termed NimbleScreen 4. In this format, the available array is divided into a 2 x 2 grid of subarrays within which up to 55 000 user-defined probes can be synthesized. The working sample volume is 10-30 ^L, and the method of sample containment is analogous to that in NimbleScreen 12. A template for four-well array design is available as an option for array layouts. The sample fixture described above is employed for four-well hybridizations, but the in-

Fig. 4-8 Nimble Screen 12 gene expression profiling. The image schown is a NimbleScreen 12 hybridization of mouse liver cRNA to a mouse liver/spleen design. Genes empirically selected for high-level expression in mouse liver were placed in the upper half of each well and genes highly expressed in spleen were placed in the lower half. The inset shows a close up view of one of the wells. The interwell r2 values for the extracted data sets had an average of 0.9972.

Fig. 4-8 Nimble Screen 12 gene expression profiling. The image schown is a NimbleScreen 12 hybridization of mouse liver cRNA to a mouse liver/spleen design. Genes empirically selected for high-level expression in mouse liver were placed in the upper half of each well and genes highly expressed in spleen were placed in the lower half. The inset shows a close up view of one of the wells. The interwell r2 values for the extracted data sets had an average of 0.9972.

terchangeable sample insert is replaced with one designed to align and contain four samples, rather than 12. The sample volume is optimized to incorporate a small bubble during sample loading. The bubble facilitates sample mixing as the fixture is rotated during hybridization, providing hybridization uniformity. The higher probe capacity of the four-well system may be well suited to the evaluation of smaller toxico-genomic training sets in which a larger number of samples is desired, for example, when working with smaller genomes or applying a training set with a reduced number of genes. With a capacity of up to 55 000, it is possible to query expression levels of 10 000 genes with a representation of five probes per gene in each of the four wells of NimbleScreen 4.

Fig. 4-9 Nimble Screen 12 gene expression profiling. The array shown incorporates probes sets derived from genes previously determined to be highly expressed in mouse liver and spleen (an expanded set analogous to Figure 4-7) tiled and interleaved. Each well was hybridized with mouse liver cRNA. Each well contains 55,000 features and includes a mismatch control for each perfect match probe.

Fig. 4-9 Nimble Screen 12 gene expression profiling. The array shown incorporates probes sets derived from genes previously determined to be highly expressed in mouse liver and spleen (an expanded set analogous to Figure 4-7) tiled and interleaved. Each well was hybridized with mouse liver cRNA. Each well contains 55,000 features and includes a mismatch control for each perfect match probe.

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