Are tumor tissue microarrays stable for FISH probe validation?

For new probe validation, it is not recommended to start with a full-size large chip. Instead, begin with 1–2 small-format chips or trial slides containing positive/negative tissues to run gradients for proteinase K concentration, denaturation temperature, and washing stringency. Since TMA places multiple cancer types on the same slide, digestion conditions must be compromised; some tissues may be over-digested or under-digested. Record signal and background differences for each cancer type, and if necessary, use parallel validation with regular sections from the same batch.

TMA section loss is mainly related to slide surface treatment, baking, and hybridization conditions. Even if the supplier has performed anti-detachment treatment, it is still recommended to bake the slides at 60-65°C for 2-4 hours before the experiment, or overnight as recommended by the kit, which can significantly reduce section loss during the denaturation step. Do not blindly increase the baking temperature or extend the baking time too long, as this may damage DNA, causing weakened signal or increased background.

Most commercially available tumor TMAs are neutral formalin-fixed paraffin-embedded with a section thickness of 4-5 μm, which is theoretically suitable for FISH. However, fixation time and dehydration protocols from different sources directly affect hybridization efficiency and signal intensity. Before purchase, confirm the fixation method and avoid tissues fixed with mercury-containing, picric acid, or heavy metal fixatives. When possible, request FISH quality control examples from that tissue batch, such as signal images for HER2 in breast cancer/gastric cancer or MET in lung cancer.

Some suppliers provide FISH compatibility QC data or empirical conditions, but most can only serve as a reference and cannot replace your own validation. You can ask them to provide the catalog numbers and probe types that have been used for FISH; if no ready data are available, at least confirm that positively charged or silanized adhesive slides are used for sectioning. Upon receipt, first check for section loss and tissue folding. If the section loss rate is high, try changing the type of adhesive slide, extending baking time, or lowering the denaturation temperature.