Procedural Overview

Cells recovered from urine samples are fixed on microscope slides. The DNA is denatured to its single stranded form and subsequently allowed to hybridize. Following hybridization, the unbound probe is removed by a series of washes, and the nuclei are counterstained with DAPI (4,6 diamidino-2-phenylindole), a DNA-specific stain that fluoresces blue. Specific hybridization of the UroVysion probes is viewed using a fluorescence microscope equipped with appropriate excitation and emission filtersets allowing visualization of the intense red, green, aqua, and gold fluorescent signals. Enumeration of the probes specific for chromosomes 3, 7, and 17, and the 9p21 region is conducted by microscopic examination of the nucleus, which is easily distinguished due to the DAPI staining.  

Bladder cells are isolated from voided urine via a series of centrifugation and fixative washing steps that remove debris and help stabilize the cells for later analysis.

Isolated and fixed bladder cells are dropped/mounted on microscope slides. A series of washes and enzymatic treatments remove proteins and other cell structures that might prevent the fluorescent DNA probes' entry into the nucleus.

Pretreated cells are heated to unwind their chromosomal DNA and allow access of the administered probe DNA(s). Subsequent cooling to a steady incubation temperature allows the probe DNA(s) to hybridize to target DNA over several hours.