cloning cylinders

Zhonglin Chai Zhonglin.Chai at med.monash.edu.au
Tue Sep 7 18:39:19 EST 1999


Tobias,

I used to have the same problem, and asked the same question in this group.
Now I have been using an alternative method: filter paper discs, and it
works great in my hand (many thanks to the people who replied and offered
me helpfull suggestions). Briefly, puncture some 3mm filter paper with 2 or
3 hole puncturer to make filter paper discs (~6mm diameter). Autoclave
them. Soak some discs in Trypsin/EDTA (TE), and using a fine end forceps
(ethanol sterilised) to carefully put a disc on the well-separated colonies
which have been washed with PBS. Leave the TE discs on colonies for ~ 5min,
and then simply use the forceps to transfer the discs to 24 well plates
which contain the desired medium. Put the discs cell side down, and the
cells will attach to the surface of the palte well, and grow. I usually
leave the discs in the wells until next medium change. Hope this of help.

Zhonglin Chai

>i'm facing serious problems trying to isolate stable mammalian cell clones
>using self-made cloning cylinders.  leaking is the big problem. are there
>any methods that are more reliable?
>
>Tobias

ZhongLin Chai, PhD
__________________________________________________________
Department of Pathology and Immunology
Monash University Medical School
Alfred Hospital
Commercial Rd, Prahran, VIC 3181, AUSTRALIA
Telephone: (61 3) 9903 0698 (lab)
           (61 3) 9903 0696 (office)
Mobile:	   0413 58 1940 or International: +61 413 58 1940
Fax:       (61 3) 9903 0731
email:     zhonglin.chai at med.monash.edu.au
___________________________________________________________





More information about the Methods mailing list