Dear all,

I am running an RNAscope experiment on human fetal spinal cord sections, roughly gestational week 15. After running my latest round of experiments, I've noticed a brown/bronze tint on my sections, pretty much everywhere DAPI (nuclei) should be. I have attached an image for reference. I've had this happen once before, but I am not too sure what is happening. I've run RNAscope many many times, and it's only happened twice. Consequently, when I image these sections under confocal, my signal (puncta) for the probes I am looking for are non-existent. I run a standard probe that I've used in all my experiments along with an exploratory probe, so at the very least, the standard probe should definitely be showing signal. This standard probe, let's say probe x, has consistently shown signal/puncta, but in these sets of sections with this strange brown/bronze hue, it's non-existent. Any ideas as to what is causing this? Or has anyone run across the same or similar issue? I am unsure how to troubleshoot this problem, I've been following the protocol very critically so unsure what happened.

Thanks so much in advance,

A