Dear Melanie, freezing-thawing hardly affects viability of lymphocytes. After this procedure they will  be unable to produce T cell responses. This is because cellular organelles and membranes are destroyed by ice needles. Even the use of crioprotectors during freezing of cells and FACS analysis immediately after thawing will result in compromised results. So, my advice is to use freshly isolated viable lymphocytes without any freezing-thawing procedure.

 Thank you Professor Kazansky. Unfortunately the cells I have available are already frozen so we are going to give it a try and see how we go. Coordinating having antigen-presenting cells and fresh tumor draining LNs available on the same day is a logistical challenge, hence the frozen LNs. After speaking with my supervisor we are going to rest them overnight. Fingers crossed that the T cells survive the process.

Dear Melanie, I would not recommend to spend your time. With frozen cells you can isolate DNA, RNA, proteins, to measure enzymatic activity, but you will never see T cell responses. If you intend to save viable (not frozen) T cells overnight, this is possible. The best way to save T cells by this mean would be placing into tightly closed flasks in complete medium and saving them in the dark at RT.