Recent advances in genomics, including serial analysis of gene expression, and DNA microarrays have allowed researchers to perform high throughput analysis of gene expression. These experiments generate large amounts of information that must be validated independently, one gene at a time. We developed an inexpensive method that works with liquid samples (tissue homogenates) and antibodies, which were developed for Western blotting but can be extended for other types of biological assays. Wells made in a frozen block of embedding material were filled with biological samples, which freeze and bond to the surrounding block. The loaded block was cut in a cryostat, and sections were transferred to nitrocellulose-coated slides for subsequent analysis. We evaluated the sensitivity and reproducibility using recombinant protein (PSA) and kidney tissue homogenates. We detected a 50% decrease in NaK-ATPase in ischemic kidney by cyroaray and by traditional western blotting. We found that cryoarrays are a moderate throughput proteomics platform that can detect both recombinant and tissue proteins. The sensitivity and reproducibility of cryoarrays is similar to other moderate and high-throughput methods8,11. The ability to make multiple (i.e., up to 800) cryosections from one cryoblock enables parallel analysis of many identical arrays. Unlike other proteomic techniques, cryoarrays are easy and inexpensive to produce, efficiently use samples with little waste, require only a small volume of sample, and are protein friendly because samples are kept frozen during production. The cryoarray method allows small laboratories without access to expensive arraying equipment to produce many identical arrays with moderate numbers of precious samples. Proteins can be detected in their native configuration, without SDS or formalin. Cryoarrays may be useful for screening small samples of precious biological fluids or tissues for new biomarkers or for rapid screening of monoclonal antibodies. It may be possible to use cryoarrays to also measure protein function and protein-protein interactions. In addition, the system could be modified to serve as a delivery device to deliver virus and bacterial samples to an organism. We also worked on developing Laser Capture Microdissection for use in kidney tissue, and patented an Immunofluorescence targeted LCM method
| Murakami, H; Liotta, L; Star, R A (2000) IF-LCM: laser capture microdissection of immunofluorescently defined cells for mRNA analysis rapid communication. Kidney Int 58:1346-53 |
| Kohda, Y; Murakami, H; Moe, O W et al. (2000) Analysis of segmental renal gene expression by laser capture microdissection. Kidney Int 57:321-31 |
