Hello,
I am doing miRNAscopes on frozen mouse retina sections (12 μm thick) adhered to Superfrost-OT Plus microscope slides. After retina sectioning, I put slides in the oven at 37 °C overnight to strengthen the tissue adherence to the slides and then store them at -20 °C.
During the miRNAscope procedure, I encountered an issue where most retinal sections lost their normal structure and became partly detached from the slide. I think this issue may arise during the retrieval step, which involves boiling the retinal sections in a specific reagent at 100 °C for approximately 15 minutes, followed by immediate immersion in cold MQ water. Does anyone have any ideas or related experience? Is there a method to improve the adhesion of sections to slides that protect them during boiling?
Thank you,
I can give you just a general hint. In the retrieval step time and temperature are related. So if you try to lower the temperature and prolong the time, it may have a benefit for adhesion. Also I think that chilling the hot slides immediatly may lead to tissue-loss. For IHC I have learned that the slides have to chill slowly - but I don't know, if this is part of the given procedure.
Hi Gudrun,
Thank you for your response. I also suspect that chilling the hot slides immediately lead to tissue detachment or damage, as it happened for me before when I used boiling citrate buffer for retrieval step in IHC.
And when I let slides chill slowly for second run of IHC, there wasn't any detachment.
But the issue is that, the RNAscope protocol says slide transferring to water is immediately. Now, I don't know if I can chill my slide a little before washing or not. I have been waiting for ACD technical support response.
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