For hard to thaw lines we always use feedercells, I most like irradiated
thymocytes about 10e6/well
On Fri, 24 Mar 1995, Mitch Magee wrote:
> > hi,
> >
> > I'm trying to resuscitate a hybridoma line that we have. I usually grow
> > the lines in DMEM HG, 10% FCS, HT, Non-Essential AA, Sodium Pyruvate,
> > HEPES. The other lines grow well in this medium but there is one in
> > particular that just won't grow.
> >
> > The line grew fine in the usual medium and was then frozen in LN2 (3-4
> > years ago; before my time). Cell viability (by Trypan Blue exclusion) was
> > fine on thawing. Now, it just won't grow in our usual medium.
> >
> > Are there any other supplements I should throw in to kick start this
> > particular line?
> >
> > I have gotten suggestions to add CEGABA, T/B cell growth factors (IL2, 4,
> > & 6), conditioned media.... etc.
> >
> > Which supplement worked the best for you? I'm running out of vials of this
> > line and I can only try a supplement or two.
>>> IL-6 has worked well for me in these situations. Something else you
> might consider is to inject the hybridoma into pristane-primed nude
> mice. In vivo conditions may provide multiple factors that cannot be
> duplicated with recombinant cytokines.
>>>>>>> Mitch Magee
> Department of Research Immunology
> Texas Center for Infectious Disease
> 2303 S.E. Military Dr.
> San Antonio, TX 78223-3597
>> Phone: (210) 534-8857 xt 209
> Fax: (210) 532-7791
> Email: MMAGEE at tcid.tdh.texas.gov>>>
