It appears that the limiting step using magnetic beads to isolate cells is
the cost of the magnet. We already have a magnet (fairly old) that takes
10ml tubes. Could this be used in the Milteyni / Stepsep systems or do you
have to buy the correct magnet? Also what is your recovery rate of T-cells
from a preparation of PBMC?
Andrew Hall
University of Aberdeen
Kenneth Frauwirth <kfrauwir at midway.uchicago.edu> wrote in message
news:HK3N4.131$v3.1856 at uchinews...
> In article <8e1iro$5f5$1 at SonOfMaze.dpo.uab.edu>,
> James F. George, Ph.D. <jgeorge at REMOVE_THISuab.edu> wrote:
> >We have made extensive use of the Miltenyi system with good results for
both
> >depletion and enrichment experiments. The beads do not seem to interfere
> >with functional assays or the ability of the cell types we are studying
to
> >mediate their function in vivo.
> >
> >The largest drawback is, if you use their complete MACS system, it
requires
> >a fairly hefty up-front expenditure because the magnet is expensive.
>> Another alternative is the StemSep system by StemCell Technologies. The
> up-front cost is less because the magnet is less expensive (although I
> think the Miltenyi magnet is easier to use), and it is a little more
> flexible because the Ab isn't directly linked to the beads. The beads are
> coated with dextran, and the antibodies are biotinylated; a bifunctional
> anti-biotin/anti-dextran antibody is then used to crosslink the two (this
> might also avoid your steric hindrance problems). This allows the use of
> a cocktail of antibodies for separation purposes, making negative
> selection purification very efficient (I routinely do T cell preps of >95%
> purity, and there are no beads bound to the T cells at the end). It also
> allows you to use your favorite antibodies without having to covalently
> link them to beads - all you need to do is biotinylate them.
>> Ken Frauwirth
> --
> Ken Frauwirth (MiSTie #33025) kfrauwir at midway.uchicago.edu> Abramson Cancer Research Institute
> University of Pennsylvania
>http://mail.med.upenn.edu/~kfrauwir
