INTERLABORATORY VARIABILITY IN FLUORESCENCE IN-SITU HYBRIDIZATION ANALYSIS

Reliable interpretation of fluorescence in situ hybridization (FISH) d ata, especially data that have been generated in more than one laborat ory, requires knowledge of the sources of variability inherent in FISH analysis, Possible sources of variation may derive from differences i n sample preparation, probes used, intrasample heterogeneity, hybridiz ation protocols, counting criteria within and between scorers, fluores cence microscopes, and filters. This study characterized the relative weight of some of these factors in order to determine the degree to wh ich FISH results are comparable between laboratories, We used a hierar chical partitioned chi(2) analysis to measure sources of variation. We found that replicate counts varied no more than expected based on cou nting statistics (i.e., multinomial variation), However, with replicat e hybridizations done in two separate laboratories, the variability in creased significantly, Thus, care must be taken when interpreting FISH data that are derived from more than one institution, Previously agre ed upon counting criteria as well as standardized FISH hybridization p rotocols may decrease this variability. (C) 1996 Wiley-Liss, Inc.