I am currently looking into reviving T-cells or B-cells from frozen human tissue. Is it feasible? If yes, is there a specific way on how to freeze the tissue in the first place to get the best possible outcome?
This is for immunotherapy-based studies, so in general how a human tissue sample can be stored?
Thank you in advance
Dear Ioannis
Yes you can isolate T/B cells from frozen tissue but they won't be viable unless they were frozen in liquid nitrogen. So if you are after cells for immunocytochemistry or immunohistochemistry, you can do it from a frozen tissue (-20-80C) but if you are after viable cells for functional assays, the tissue should have been stored in correct conditions. Hope that answers your question. Good luck. Siva
Hi,
Please check the link below. This report may contain helpful information about your question.
Best wishes SM Najim
http://www.sciencedirect.com/science/article/pii/S1658387608500409
Dear
I recommend you an interesting mansucript entitled: Reconstitution of paired T cell receptor α- and β-chains from microdissected single cells of human inflammatory tissues. Proc Natl Acad Sci U S A. 2006 Aug 8; 103(32): 12057–12062.
Regards
Marius
A visit to this site may be of help:https://www.stemcell.com/.../BRDX20366-Tools_Immunology_Research.
For tissue storage (cells suspension) resuspend cells in freshly prepared freezing solution (10% DMSO, 20% human serum, PBS or corresponding cells medium) with maximum cells concentration 50-100 million cells/ml and distribute in cryovials (properly labelled in advance). Put vials in freezing-box (usually filled with isopropanol and cooled in advance to 4 C in the refrigerator). Operate as quick as possible since DMSO is toxic for cells. Immediately transfer freezing boxes at -80C for at least 24h but no longer than one week. Afterwards, transfer cells into liquid nitrogen tank (-150C or -180C) for long-term storage and do not touch them for at least one week to let them settle down. Write down the exact location of your cells ;D
To thaw cells: use warm (37C) medium 10-30 ml, containing 10% human serum. Put vials in the water bath to get ice melt (not completely, appx 50%) and then transfer cells with unmelted ice into the tube with the warm medium. Mix carefully, since unmelted ice crystals can damage cells from inside. Centrifuge cells and remove supernatant, resuspend cells in PBS and centrifuge cells again. After this second washing, you can count them. Because if you count them immediately - you get wrong cell numbers since many cells die, but it takes some time. Normally 1/3 of cells die after freezing-thawing cycle.
Good luck!
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