[Cytometry] CS&T

Felix Heymann felix.reichel at ukb.uni-bonn.de
Tue Jul 21 02:08:17 EDT 2009

Hi Cris, dear all,
thank you for the addition. The problem with the instrument 
configuration really is, that whatever has been changed by the last user 
stays on the machine as far as I got it, am I correct here? So actually 
for my regular flow-users I never talk about laser delay setting, area 
scaling and window extensions, because on the analyzers they simply do 
not need to touch it anyway normally (or otherwise if they do, they will 
be beheaded, first by me and by the users with messed up experiments 
afterwards....), but for the sorters of course these values may have to 
be adjusted. One more thing about the CS&T that troubled me already for 
some time I want to ask and see if some else out there has the same 
issues. If I run the baseline and performance I get decent results in 
terms of the rCVs and SDs, but for some channels the machine totally 
over-(or under-)estimates the PMT voltage settings (I just did a run 
yesterday and ended up e.g. with a PMT of 417V for my PerCPCy5.5 amp, 
which is on our machine rediculously low). This is of course a problem 
when it comes to real settings, so I normally readjust the values again 
anyway and very rarely stick to the CS&Ts. Can anyone explain to me why 
that is? I like the idea behind the CS&Ts, but for me it seems that 
there are either some mishandlings or some problems which I am not fully 
aware of.

Thanks for your advide


Felix Heymann
Scientific research fellow
Medical clinic III, University hospital Aachen

Christopher Bare schrieb:
> Felix, Neal, Pam,
> Please be warned that using CS&T in a multi-account (i.e., multiple login
> names inside DiVa) can have drastic effects on laser timing and delay if
> there are mismatches in several database elements: Instrument Configuration,
> Current CS&T settings, and last tube acquired by user.
> It is also possible for users in a multi-account system to change laser
> setup values EVEN IF they do not have permissions as assigned in the
> "Adminstration" window.
> -Cris Bare
> Rockefeller University
>> -----Original Message-----
>> From: cytometry-bounces at lists.purdue.edu
> [mailto:cytometry-bounces at lists.purdue.edu]
>> On Behalf Of Felix Heymann
>> Sent: Friday, July 17, 2009 7:11 PM
>> Cc: Cytometry List
>> Subject: Re: [Cytometry] CS&T
>> Hi Pam,
>> I just came back from an Aria II training from BD today (lucky me, the
>> guys did a real terrific job!) and of course we did a lot of baseline
>> configs and performance checks to calibrate the instruments. Our
>> trainers actually stressed that the bead lot no is quite important,
>> since there are quite some informations (I guess mostly about the actual
>> fluorescent properties of the specific lot)  hidden in the lot files
>> sitting on your instrument. If there's new bead lots out which have not
>> been listed on your instrument, you should actually also download the
>> actual information from the BD website. Since the machine uses the CS&T
>> to set also your laser delays  and area scalings, not only the PMTs,
>> working with the wrong lot no actually could affect the accurate digital
>> allignment of your instrument, so from my point of view it's quite
>> necessary to stick to the lot no if you run a performance check
>> belonging to a certain baseline. If you do regular performance checks,
>> you can also keep track about the development of your intrument
>> performance and can see early if e.g. a laser starts to get bad. Since
>> core facilities have a high throughput of external customers, regular
>> quality controls guarantee a high level of performance from your core
>> and keep customers happy......and in case things go wrong you can at
>> least prove that it was not due to a problem with the machines.
>> best
>> Felix
>>> Hi Pam,
>>> We have multiple "configurations" of the instrument with the different
>>> bead lots.  I ran separate baselines for the users.  The user just has
>>> to make sure to pick the right configuration and can do the
>>> performance checks as needed.  Please let me know if you need any
>>> further details.
>>> Good luck,
>>> Neal
>>> ICBR Flow Cytometry Facility
>>> University of Florida
>>> On Jul 6, 2009, at 2:10 PM, Moody, Pamela wrote:
>>>> Hi Everyone,
>>>> Our core facility has an Aria I, Aria II and LSRII which are all
>>>> running
>>>> CS&T beads.  We have trained users which are allowed to run these
>>>> instruments and run the CS&T beads.  Each of these users has a
>>>> different
>>>> lot number of CS&T beads.  I would like to know what other core
>>>> facilities do in this situation.  Can you run different bead lots with
>>>> the same baseline?  Do you ignore the bead lot?  New users need to do
>>>> everything by the book and exactly how they were shown in class.  They
>>>> don't always know what it means if you ask them but they want to do it
>>>> anyway.  I could go on forever about this but I would rather hear your
>>>> stories.
>>>> Thanks
>>>> Pam
>>>> Pamela Moody
>>>> Manager, Flow Cytometry Shared Resource
>>>> Cold Spring Harbor Laboratory
>>>> Hershey Building
>>>> One Bungtown Road
>>>> Cold Spring Harbor, New York 11724
>>>> Phone: (516) 367-8477
>>>> Fax: (516) 367-8461
>>>> Website: http://www.cshl.edu/cancercenter/shared_resources.html
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