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Fig 2 (FISH guide)
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Legend Fig. 2. a-f) Trypsin pretreatment. a-c) Slides kept 1-2 hours at RT only, were subjected to trypsin pretreatment for 15 seconds (a) and 6 seconds (b) or no trypsin (c), then denatured and DAPI stained. Increased trypsin treatment altered chromosome morphology. d-f) Chromosome 1 centromere was hybridized onto non-pretreated (d), 10 second pretreated (e) and 5 seconds pretreated (f) slides. FISH r esults are better on the pretreated slides. Arrows point to chromosome 1 signals. g-i) Post-denaturing trypsin treatment onto chemically aged, less optimal cell suspension. Slides prepared from a less than optimal cell suspension were subjected chemical aging for 10 seconds (2g), 50 seconds (2h) or 2 minutes (2i), followed by denaturing and G-banding. Results improved somewhat with longer aging, but were worse overall than with a better cell suspension (2k). j-l) Chemical aging vs. dry heat on optimal cell suspension. Slides prepared from an optimally prepared cell suspension were subjected to chemical aging 2 minutes and then were either left non-denatured (2j) or were denatured (2k), or were subjected to 94 C dry heat 1 hour, were not denatured (2l) and all slides were stained with Giemsa. Results show that dry heat is clearly better than chemical aging for G-banding, and that denaturing is detrimental to banding quality. A clean, well prepared cell suspension produces better bands than a lower quality suspension (2k vs. 2i).
