isolation of nuclei

Administrador de iihema.fmed.uba.ar postmast at iihema.fmed.uba.ar
Fri Mar 15 23:41:27 EST 1996


Dear Kristina Warton <K.Warton at cfi.unsw.EDU.AU>:

You wrote:

>I'm looking for a protocol for isolating nuclei from cultured human 
>and rodent cells. I'm after a protocol which does not use detergents, 
>so that the membranes of the nuclear envelope remain intact. The 
>protocols I have been using either leave most of the cells 
>undisrupted, or cause the nuclei to clump. I'm worried that spinning 
>clumps of nuclei through sucrose to purify them will not do anything 
>towards removing the cell debris around individual nulcei.

I think the following papers may have the information you need:

"Enucleation of mammalian cells by cytochalasin B" (I and II).(Poste, G.
& Reeve, P.)
I = Exp. Cell Res. 73:273. 1972.
II= Exp. Cell Res. 73:287. 1972.

They just culture the cells with cytochalasin B, spin them and voila!, obtain
enucleated cells.
You may also find a more recent usage of these techniques in:
"Cell nucleus and DNA fragmentation are not required for apoptosis",
Journal of Cell Biology 127:15 (1994).
I must say I have never used these protocols yet, so I can't assure they 
work.  Anyway intend to apply these techniques in the near future, so I'd
really appreciate some feedback if you ever get to try them.

Hope it helps you.  Good Luck!!.

Facundo Garcia Bournissen,
Academia Nacional de Medicina,
Inmunologia Oncologica,
Pacheco de Melo 3081,
Buenos Aires, Argentina.

Email= postmast at iihema.fmed.uba.ar
Fax= (541) 803 9475




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