[Cytometry] FW: flow cytometry question

christopher.fogarty at helsinki.fi christopher.fogarty at helsinki.fi
Tue Feb 4 02:18:29 EST 2014

Hi Pam,

I have previously worked with bacteria in the FACS machine and have  
found a pretty significant change in scatter properties if I don't  
vortex the samples just before the run. Given that it's relatively  
straight forward to see the different scatter properties before and  
after vortex, it very well may be possible to see differences in  
clumping between a population that clumps and one that doesn't as much  
using FSC-A vs -H and SSC.

It seems that testing the hypothesis would be pretty straight forward,  
as least. But keep in mind that vortexing affects the scatter  

Hope this helps,


Quoting "Shaw, Pam" <pam.shaw at mso.umt.edu>:

> Hello flow list buddies!
> I have a request a bit out of my  comfort zone.  We don’t do a lot  
> of bacterial work, so could use some input on this request (see  
> below).  We have a FACSAria II with 405, 488 and 633 nm lasers.   
> Would the 633 laser work for cyanobacteria as described below and  
> would we be able to see a difference in size as the request indicates?
> Any input would be appreciated!
> Thanks in advance.
> --
> Pam Shaw
> Fluorescence Cytometry Core Facilitator
> Center for Environmental Health Sciences
> The University of Montana
> 32 Campus Dr, Skaggs Building 052
> Missoula, MT 59812
> Ph (406) 243-4974
> Email:  
> pamela.shaw at umontana.edu<applewebdata://DE30ADE2-B1DF-44AF-B224-B0B84EDBD782/pamela.shaw@umontana.edu>
> www.umt.edu/cehs/Facility_Cores/Fluorescence_Cytometry/default.aspx
> From: <Miller>, Scott  
> <Scott.Miller at mso.umt.edu<mailto:Scott.Miller at mso.umt.edu>>
> Date: Friday, January 31, 2014 at 12:51 PM
> To: "pamela.shaw at umontana.edu<mailto:pamela.shaw at umontana.edu>"  
> <pamela.shaw at umontana.edu<mailto:pamela.shaw at umontana.edu>>
> Subject: flow cytometry question
> Hi Pam,
> By way of introduction, my name is Scott Miller, a microbiologist in  
> DBS, with a research question that may potentially be appropriate  
> for flow cytometry. The gist is that we are comparing wild type and  
> mutant cyanobacteria in one of our projects. Under certain  
> conditions, the mutant appears “clumpier” than the wild type,  
> probably because of differences in extracellular polysaccharides,  
> and we are exploring possible methods for quantifying this. These  
> bacteria autofluoresce if excited with ~590 nm (peak emission at 680  
> nm), and so we were curious whether it might be feasible to obtain  
> particle size distributions between wild type and mutant by flow  
> cytometry. The bacteria are filamentous and the clumps are very  
> small by macroscopic standards, but I don’t know how this lack of  
> homogeneity of the sample would impact the method. Many thanks for  
> any advice that you might have and for your thoughts on the  
> feasibility of the approach for this question.
> Cheers,
> Scott
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