Arrin Katz

Nanoliter high throughput quantitative PCR

Understanding biological complexity arising from patterns of gene expression requires accurate and precise measurement of RNA levels across large numbers of genes simultaneously. Real time PCR (RT-PCR) in a microtiter plate is the preferred method for quantitative transcriptional analysis but scaling RT-PCR to higher throughputs in this fluidic format is intrinsically limited by cost and logistic considerations. Hybridization microarrays measure the transcription of many thousands of genes simultaneously yet are limited by low sensitivity, dynamic range, accuracy and sample throughput. The hybrid approach described here combines the superior accuracy, precision and dynamic range of RT-PCR with the parallelism of a microarray in an array of 3072 real time, 33 nl polymerase chain reactions (RT-PCRs) the size of a microscope slide. RT-PCR is demonstrated with an accuracy and precision equivalent to the same assay in a 384-well microplate but in a 64-fold smaller reaction volume, a 24-fold higher analytical throughput and a workflow compatible with standard microplate protocols.

Massively parallel microfluidics platform for storage and ultra-high-throughput screening

We have developed a novel microarray technology for performing very large numbers of biochemical, chemical and cell-based nanoliter volume synthesis, storage and screening operations in a massively parallel manner. The Living Chip is an array of precisely machined through-holes retaining nanoliters of fluid by capillary action. Sample loading, washing and recovery are operations that can be performed manually or with simple automation. Mixing between co- registered through-holes is achieved by stacking two or more precision aligned arrays and optical assay read-outs are in parallel with a CCD imaging system. An automated picker system transfers hits into lower density microtiter plates for further analysis. We will present result demonstrating massively parallel implementation of both homogeneous and inhomogeneous fluidic and cell-based assay systems and applications of the chip for drug compound library storage and management.

Antibody Library Screening in Through-Hole Microarrays

Through application of recombinant DNA technology monoclonal human antibody libraries of over 109 antibodies are stored as plasmids in bacterial (E. coli) colonies. We have developed ultra high throughput methods suitable for screening these libraries after only one round of panning in precisely constructed, high density arrays of micro-channels, or bottomless wells.