[Cytometry] Offbeat ergonomics question

Sasha Lazetic Sasha.Lazetic at oncomed.com
Thu Mar 4 17:13:36 EST 2010


Hi Cynthia,

We run up to 16 96 well plates a day and we also run 384 well plates. For the washing steps we dump our sups or wash buffer and vortex the plate when it is 'empty' at top speed 2-3 pulses. There is always 10-20ul of residual volume in the well and that is just fine for re-suspending a cell pellet without splashing cells into adjacent wells. Adding 250ul wash buffer with a multichannel or an ELISA plate washer to a pellet that has already been re-suspended by vortexing re-suspends the pellet sufficiently.

For addition of antibody reagents and subsequent incubation, we again vortex the plate before the addition of the antibody (usually around 50ul, but as much as 200ul for sups). We also resuspend the cells on an ELISA plate shaker briefly after the addition of the antibody. For 50ul volumes we find top speed on the plate shaker turned on for a few seconds is good. For 200ul in a 350ul plate, the speed of the shaking needs to be ramped up by hand a little more slowly than for 50ul, but works just as well. RSI is no fun!

Good luck,
Sasha Lazetic
OMPI

-----Original Message-----
From: cytometry-bounces at lists.purdue.edu [mailto:cytometry-bounces at lists.purdue.edu] On Behalf Of Pieter Bogaert
Sent: Wednesday, March 03, 2010 7:18 AM
To: Mara Rocchi
Cc: Cytometry at lists.purdue.edu
Subject: Re: [Cytometry] Offbeat ergonomics question

maybe this is too simple, but what about resuspending with a 
multichannel pipet? We do that all the time... When having a lot of 
plates, you use a lot of tips, but filling a box takes about 3 minutes 
if they don't have to be sterile...
grts
Pieter


Op 3/03/2010 15:20, Mara Rocchi schreef:
> Dear Cynthia,
> I assume here we are talking of cells stained in 96-well plates for FACS.
> After centrifugation, I vortex the plate very gently (after discarding the supernatant) enough to resuspend the pellet. To avoid cross-contamination I use only alternate wells, both horizontally and vertically.
>
> Hope this help
> Mara
>
> Mara Rocchi BVM&S, PhD
> Flow Cytometry Manager
> Moredun Research Institute
> Pentlands Science Park
> Bush Loan, Penicuik
> EH260PZ
> Scotland, UK
>
>
> Think before you print. Save paper and help the environment-----Original Message-----
> From: cytometry-bounces at lists.purdue.edu [mailto:cytometry-bounces at lists.purdue.edu] On Behalf Of Cynthia Martino
> Sent: 02 March 2010 15:05
> To: Cytometry at lists.purdue.edu
> Subject: [Cytometry] Offbeat ergonomics question
>
> OK, so I've been working in the lab long enough to have developed a nasty case of tendinitis.  Does anyone out there have an ergonomic alternative to resuspending cells in a plate as opposed to banging on each side?
>
> I realize this is not typical of the topics discusses on this forum but thought it might be the fastest/easiest way to find a less painful alternative.
>
> Thanks!
>
>
>
>
>
> Cynthia Martino
> Senior Research Assistant
> Cooper Lab
> Trudeau Institute
> 154 Algonquin Ave.
> Saranac Lake, NY  12983
> (518) 891-3080 X-145
> cmartino at trudeauinstitute.org
>
>
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-- 
Pieter Bogaert
Flow cytometry core facility
Ghent University&  VIB
Fiers-Schell-Van Montagu building
Technologiepark 927
B-9052 Ghent (Zwijnaarde), Belgium

Tel: +32-9-33 13 642
E-mail: pieter.bogaert at dmbr.vib-UGent.be

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