[Cytometry] What are the 4 peaks on our cell cycle analysis

W.E.Corver@lumc.nl W.E.Corver at lumc.nl
Thu Feb 18 03:57:27 EST 2010


Dear Daniel,

Since you are performing a relative measurement you can only determine
DNA aneuploidy by flow using a reference. The peaks at 150 are very
likely clumps. A triploid population (3n) would pop-up at 75. At 200
might also represent clumps. However, many cell lines in culture undergo
endo-reduplication, meaning that G2 cells at 100 do not enter mitosis
but start a second cycle of DNA synthesis. Suppose you have 2n cells to
start with (at 50) the peak at 200 might than also represent 8n cells or
be a mixture of clumps and 8n cells.

For DNA content analysis I advise you to use sophisticated software like
ModFit or MultiCycle. These software packages include algorithms to
correct for debris, clumps / aggregates, etc.

Send me your file and I will analyze it for you.

Kind regards,

Willem Corver


Willem E. Corver, PhD
 
Dept. of Pathology
Leiden University Medical Centre
P.O. Box 9600, Building 1, L1-Q
2300 RC  Leiden
The Netherlands
 
Tel: +31 71 526 6580
FAX: +31 71 524 8158

-----Original Message-----
From: cytometry-bounces at lists.purdue.edu
[mailto:cytometry-bounces at lists.purdue.edu] On Behalf Of Daniel,
Benjamin J
Sent: donderdag 18 februari 2010 3:20
To: cytometry at lists.purdue.edu
Subject: [Cytometry] What are the 4 peaks on our cell cycle analysis

Dear all,

I am new to cell cycle analysis. I have a tumor cell line that shows
nice tight peaks at 50 and 100 that represent G0G1 and G2M respectively.
I have some smaller peaks at 150 and 200.  We believe that this tumor
line is aneuploidy. This is where I get lost. Would the peak at 150 be
triploidy G0G1 and the peak at 200 be aneuploidy G2M?

I've performed the doublet discrimination as suggested by every protocol
that i've come across. and I believe these last two peeks are singlets.
Since I"m new to this, I may be wrong. Could the peak at 150 be doublets
of cells from the G0G1 and G2M and the 200 peak be doublets of G2M?

I've seen several histograms of aneuploidy populations and this
populations seems to be all over the place.

Any suggestions would be greatly appreciated,

Ben

Benjamin J. Daniel, Ph.D.
Flow Cytometry Core Assistant Director
Instructor of Medicine/Research
Univ. of Texas HSC at San Antonio
Office of the Vice President for Research
7703 Floyd Curl Drive, Rm 5.044V
Mail Code 7763
San Antonio, TX 78229
Office Ph: (210) 567-3911
Email: danielb at uthscsa.edu<mailto:danielb at uthscsa.edu>

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