I did liver MNCs isolation for several times and I could not detect NKT cells by using NK1.1 & TCRb or α-GalCer-loaded CD1d tetramer&TCRb. During all these experiments, I used 40μm cell strainer, which was not commonly used in this isolation. Common protocol suggest 200-gauge steel mesh or 100μm nylon mesh. Is this the reason? But since lymphocytes size is less than 20μm, I do not think this is the problem. Does anyone know the reason?
Hello
kindly look at link here :
Methods for detection, isolation and culture of mouse and human invariant NKT cells.
Article Methods for detection, isolation and culture of mouse and hu...
Hope help you .
Good Luck
Hello Zhengzuo.
Your tissue processing may be a problem and you need some of capillaries cut through the liver tissue plus released NKT cells. The sieve recommendation may target the NKT cells within the cut-through cappilaries to maximize recovery. Thus rare-released cells may be lost during the procedure, and you also excluded the cut-through capillaries containing abundant NKT cells.
Have a look at an article.
http://journals.plos.org/plosone/article/file?id=10.1371/journal.pone.0010288&type=printable
Best wishes.