Resuscitating a Hybridoma line

JAF at UNCVX1.OIT.UNC.EDU JAF at UNCVX1.OIT.UNC.EDU
Fri Mar 24 08:00:51 EST 1995


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
>         
> 
> 



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