Flow Cytometric Analysis of Genotoxicity

Period of Performance: 05/23/2003 - 03/31/2006


Phase 2 SBIR

Recipient Firm

Cytomation Gtx, Inc.
Fort Collins, CO 80525
Principal Investigator


The overall objective of this proposal is to develop a flow cytometry- based assay utilizing optimized flow cytometry instrumentation and cultured mammalian cells that can be routinely employed by pharmaceutical and other companies to measure the mutagenic potential of chemicals and drugs. The assay is based on a Chinese hamster ovary- human hybrid cell line, CHO AL, which contains human chromosome 11. The principal target for mutation is the gene for the CD59 antigen which is encoded on chromosome 11. The hypotheses are(1) that mutations in chromosome 11 can be detected accurately, sensitively and rapidly by flow cytometry measurement of the loss or presence of surface antigens encoded by chromosome 11 and (2) that this mutation assay system can be developed so that it could be used routinely to determine the genotoxicity of physical and chemical agents. Specific Aim 1 is to build a relatively inexpensive benchtop flow cytometer optimized for analysis of antigen loss in mammalian cells. Specific Aim 2 is to validate the flow cytometry assay for detecting mutations in CHO AL cells by measuring the mutation dose response from a panel of 25 physical and chemical agents at a range of doses. Independent studies will be done at an off-site laboratory. Specific Aim 3 is to extend the assay to include the analysis of intragenic and chromosomal mutations by measuring the presence or absence of multiple antibodies bound to different cell surface antigens encoded by separate genes on human chromosome 11. The development of this assay system would allow for rapid, accurate, relatively cheap but informative screening of pharmaceuticals for genotoxicity and substantially reduce the cost of carrying out mutation analysis.