Validation of fluorescence in situ hybridization using an analyte-specific reagent for detection of abnormalities involving the mixed lineage leukemia gene.

CONTEXT: Fluorescence in situ hybridization (FISH) is a molecular cytogenetic assay that is commonly used in laboratory medicine. Most FISH assays are not approved by the US Food and Drug Administration but instead are laboratory-developed tests that use analyte-specific reagents. Although several guidelines exist for validation of FISH assays, few specific examples of FISH test validations are available in the literature.
OBJECTIVE: To provide an example of how a FISH assay, using an analyte-specific reagent probe, may be validated in a clinical laboratory.
DESIGN: We describe the approach used by an individual laboratory for validation of a FISH assay for mixed lineage leukemia (MLL) gene.
RESULTS: Specific validation data are provided illustrating how initial assay performance characteristics in a FISH assay for MLL may be established.
CONCLUSIONS: Protocols for initial validation of FISH assays may vary between laboratories. However, all laboratories must establish several defined performance specifications prior to implementation of FISH assays for clinical use. We describe an approach used for assessing performance specifications and validation of an analyte-specific reagent FISH assay using probes for MLL rearrangement in interphase nuclei.