Our lab has flash-frozen lung tissue from multiple time points from a now-completed study. We would like to conduct flow cytometric analysis of immune populations in the lung tissue, if possible. I recognize that viability will be a nontrivial issue, but has anyone had success with flow analysis of snap-frozen lung tissue? We also have OCT frozen and FFPE blocks, if either of these would be better for flow analysis. Thanks!
Although Flow cytometry is routinely performed on frozen cells, these are usually single cell suspensions made from fresh tissues and then frozen carefully in specialized freezing media. Even then, the results (immune cell subtyping) are never as good as if you are working with fresh cells.
Frozen whole tissues are not ideal for flow cytometry. Your best bet would be to do immunohistochemistry. Regardless of how you froze the tissues, chances are you would have a lot of cell death upon thawing. If you froze them with OCT or if they are embedded in blocks, that would make flow cytometry more difficult. It would be hard to stain and identify relatively small frequencies of leukocytes that reside in the lung, unless you make single cell suspensions from fresh lung tissues. See the thread linked below for more on this topic.
https://www.researchgate.net/post/How_to_prepare_sample_for_FACS_using_Frozen_Tissue
Your best option is probably IF on the OCT embedded tissues. You could multiplex and do serial sections if you have many markers you want to look at.
- Badges
- Science method