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Dynabeads clogging the tip of the injector?

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benmoleskin

Participant

Posts: 2

Joined: Wed Sep 21, 2016 1:49 pm

Post Fri Feb 03, 2017 3:35 pm

Dynabeads clogging the tip of the injector?

Hello All,

I have been using Dynabeads (Human T-Activator CD3/CD28) to stimulate T cells and been running them through a Helios system. We were under the impression that they would be compatible with CyTOF. Unfortunately, we had clogging problems at the level of the injector with a build-up of suspicious reddy/brown coloured residue at the very tip. This residue is quite tough and difficult to remove from the tip. We are concerned that this might be due to Dynabeads. I am stimulating T cells at a 1:1 ratio, 1 x 106 T cells at a time, per sample.

Has anyone had any experience running Dynabeads through the CyTOF, specifically has anyone encountered this type of clogging?

Thanks,

Ben.
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noelcasey

Participant

Posts: 8

Joined: Fri Apr 08, 2016 8:00 pm

Post Sun Feb 05, 2017 3:55 pm

Re: Dynabeads clogging the tip of the injector?

Ben,
To help. I have some questions.
What are the size of the dynabeads when compared to Fluidigm beads?
Do you notice drift in your tune solution over a course of a day or two.
What is your cleaning procedure for the sample introduction system after you finish for the day?

The narrow bore injector in the Helios is more prone to clogging so a more regular and/or aggressive cleaning may be required. Having a couple of clean injectors as spares and ready to swap out the old one is always a good idea.

Best,

Noel
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AdeebR

Grand master

Posts: 169

Joined: Thu Mar 13, 2014 5:58 pm

Location: NYC

Post Mon Feb 06, 2017 3:12 pm

Re: Dynabeads clogging the tip of the injector?

Hi Ben,

I always remove simulator Dynabeads before running samples through the CyTOF. Just place the tube on a magnet and wash a few times with an EDTA-containing buffer and you should be able to remove the majority of the beads from your prep.

Miltenyi MACs beads are only ~50nm in diameter so likely not a problem to introduce into the instrument, but I believe Dynabeads are ~5micron. While this is still smaller than a typically cell it's still quite large, and I would imagine that they may also form aggregates with your cells which might get fixed during your cell prep. Plus, it's just a lot of extra unnecessary metal to be introducing into your system.

Adeeb
Adeeb Rahman
Icahn School of Medicine at Mount Sinai, NYC
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DaveGPPG

Participant

Posts: 1

Joined: Thu May 08, 2014 9:08 pm

Post Tue Feb 07, 2017 1:56 am

Re: Dynabeads clogging the tip of the injector?

We do not have a CyToF, but we did have an issue trying to use a bead based T-cell stimulation in an automated setting for high throughput flow cytometry. We found the Immunocult reagent from Stem Cell Technologies to be a satisfactory bead free solution for us.
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benmoleskin

Participant

Posts: 2

Joined: Wed Sep 21, 2016 1:49 pm

Post Tue Feb 07, 2017 10:42 am

Re: Dynabeads clogging the tip of the injector?

The injector cleaned up nicely.

Given that the injector blocked after running only 4 samples with beads we've decided to remove them (or as many of them as possible) before running samples. I removed the beads from the prep with a 96 well side magnet and pipetting up and down a few times.

Thanks for your help,

Ben.
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mleipold

Guru

Posts: 5796

Joined: Fri Nov 01, 2013 5:30 pm

Location: Stanford HIMC, CA, USA

Post Tue Feb 07, 2017 6:12 pm

Re: Dynabeads clogging the tip of the injector?

Hi Dave,

Thanks for that info, I know a lot of people are always looking for improved ways to stimulate their cells.

To clarify: there are several different Immunocult reagents from StemCell: Treg, Th1, Th2, Dendritic, XF Tcell, CD3/CD28, and CD3/CD28/CD2.....which one did you use? And how long did you stim?


Thanks,
Mike

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