I'm planning to isolate HIV from autopsy brain tissue in PBMC's (CD8-),
and I'm faced with several issues that I'd appreciate some comments on.
1. The samples were most-likely not handled with sterile technique at
autopsy, so I am planning to rinse the tissue with Fungizone
(anti-mycotic) and PPLO agent (anti-mycoplasma) prior to culture?
Should I continue to use Fungizone in the culture media (with
Pen/Strep/Gent) during virus isolation? What are the potential toxicity
problems (if any) with these agents, and will they affect virus
2. What is the best method for dissociating the tissue, dounce
homogenizing, finely mincing the tissue with fine scissors, repeated
motions through 23 guage cannula?? Can any method potentially destroy
3. Is anyone currently doing this with consistent isolation of virus?
I'd be very grateful for any current protocols that are working well.
Thanks in advance.
gorry at mbcrr.harvard.edu