Erythroid differentiation induction
Stephen C. Dahl
stebby at welchlink.welch.jhu.edu
Wed Dec 2 16:12:25 EST 1998
Lubomira Chakalova (lchak at OBZOR.BIO21.BAS.BG) wrote:
: Dear colleagues,
: I have just obtained a murine erythroleukemia cell line, BB88 from ATCC.
: According to ATCC erythroid differentiation and hemoglobin synthesis are
: inducible by DMSO. I have been trying to induce erythroid differentiation
: by 5-day treatment with 1.5% and 2% DMSO. I used the benzidine assay to
: detect the hemoglobin that was supposed to accumulate in at least a
: subpopulation of the cells. This assay gave negative results. There were
: cells that continued to divide up to the 5th day, while a substantial part
: of the cells died by that time.
: If anybody else has had similar problems with inducing erythroid
: differentiation, please give me some hints.
This may or may not be helpful for you...
First, be aware that in any given population of MEL cells there are those
that will differentiate well and those that won't. For this reason, some
scientists have been known to plate out replicate 96 well dishes of limit
dilution cell clones, induce one plate and clone and freeze those which
turn visably pink when induced for a few days. This is most easily seen
in cells pelleted in PBS, but you can see it in the dish too if you know
what you are looking for.
Second, MEL cells get fairly fragile as they differentiate. The dead ones
you see may have already "been there, done that". Test your induction
with treated and untreated cultures at 24 and 48 hours via benzidine and
see if you don't have a better feel for you induction rates.
Third, some cells respond better to HMBA
(N,N'-hexamethylenebis(acetamide)) rather than DMSO. Try HMBA at 3mM or
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