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THE FISH assay resulted in high slide backgroundWas slide cleaned properly prior to sample preparation? -Immerse slide in ethanol, dry using lint-free paper Were the Specimen slides used too soon prior to denaturation? -Ensure slides are aged at least 24 hours at ambient temperature Could there have been too much cellular debris on the sample? -Wash cell pellet with fresh fixative three times prior to dropping slides Were the metaphase spreads aged by baking, or do they contain cytoplasm? -Increase time the slide is immersed in the denaturation solution to 10 minutes Was the slide inadequately washed following hybridization? -Ensure wash solutions were made according to procedures outlined by the package insert -Make sure the wash solution is at the proper PH and temperature, remove cover slip and repeat the wash procedure -Increase immerse time to 4 minutes in the 73°C 0.4XSSC/0.3% NP-40 wash Were the wash solutions used too long or improperly stored? -Ensure wash solutions containing formamide are properly stored at 4°C -Discard these solutions after 7 days or after frequent use; discard all other solutions after 1 day -Ensure the pH of formamide solutions are pH 7.0-8.0 Was the hybridization viewed using long bandpass filters? - Reduce background light by switching to filters with smaller bandwidths or to multi-bandpass filters |