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Barium contamination cleanup

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JamesW

Contributor

Posts: 37

Joined: Tue Nov 21, 2017 6:59 am

Post Fri Jul 20, 2018 2:54 am

Barium contamination cleanup

Hello to all, recently we have had some problems with some human gut samples heavily contaminated with Barium. These cause persistent high barium readings after switching back to MQ water.

Can I check how others handle contamination? Currently, we stop running the sample, run MQ water and check the MPR value. If it is over 5 million intensity we turn off the plasma and run cycles of wash solution and water with the nebuliser placed into an empty tube. We also clean the cones etc. This process works but takes a long time if the initial contamination is high.

I have seen some suggestions on this forum for using Nitric acid but our local Fluidigm rep seemed quite worried when I suggested running 3% nitric acid so I wanted to ask what concentrations other people use and if it can only be done with the plasma off. I noticed that the tuning solution appears to cause a large wash out of barium which I presume is related to its nitric acid content?

We have only been running the Helios for 6 months so would appreciate some advice. We are also now asking users to screen gut samples by running 1/1000, 1/100 and 1/10 dilutions before use.

Best,
James
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ChristophS

Contributor

Posts: 21

Joined: Thu May 07, 2015 2:11 pm

Location: Switzerland

Post Fri Jul 20, 2018 12:39 pm

Re: Barium contamination cleanup

Hi James,

for the Barium contamination itself I can not say much, but for the Nitric Acid yes.
We quite routinely run 2% HNO3 (the Sigma ICP rinse solution) on our CyTOF 2.1 (Helios upgrade) with no noticable undesired effects.
This we use to clean alternatively to the Fluidigm wash (0.02% HF) since it is better in removing biological residues and not just metals.
Also we run metal content determination of newly MaxPar tagged Abs in this solution (at 1:100000 dilution).
So I would say you need not worry about HNO3 (and should tell the rep the same ;-) ).

Best

Christoph
Christoph Schwärzler
Director Cytometry
Flow Cytometry Facility (https://www.cytometry.uzh.ch/en.html)
University Zürich
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nkhanbham

Master

Posts: 53

Joined: Wed Feb 25, 2015 3:03 pm

Post Fri Jul 20, 2018 2:22 pm

Re: Barium contamination cleanup

Hi Christoph,
What is the Sigma product number for nitric acid that you use? I would like to try out your method.
Thanks,
Naeem
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ChristophS

Contributor

Posts: 21

Joined: Thu May 07, 2015 2:11 pm

Location: Switzerland

Post Fri Jul 20, 2018 2:40 pm

Re: Barium contamination cleanup

It used to be 48871 but I am afraid that is discontinued (at least in Europe).
Need to check when I am back in office from 6th August.

Best

Christoph
Christoph Schwärzler
Director Cytometry
Flow Cytometry Facility (https://www.cytometry.uzh.ch/en.html)
University Zürich
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mleipold

Guru

Posts: 5792

Joined: Fri Nov 01, 2013 5:30 pm

Location: Stanford HIMC, CA, USA

Post Fri Jul 20, 2018 3:36 pm

Re: Barium contamination cleanup

Hi all,

We have been running 3% nitric acid through all versions of our CyTOFs for close to 7 years now. As James said, Tuning solution is 2% nitric, so it's obviously ok with the fluidics of the instrument.

We dilute concentrated nitric to 3% v/v. We used Optima Grade Nitric Acid, Fisher # A467-500; Optima grade acids are certified for trace metal assays and ICP-MS, so they're super-clean.


Yes, in our experience, Nitric does a better job removing +2 metals (Ba, Cd, Pd, Pt) and Iodine than Wash solution (HF) does. Additionally, as James mentioned, just running Tuning solution will work: not only is there 2% nitric, but also other metal ions to compete with Ba (or any other contaminant) for non-specific binding sites in the PSI, tubing, etc. This can be useful if someone overtiters an antibody or has massive specific contamination/debris like CD56 debris in brain tissue, and it streaks like a tire track. Otherwise, you'll be running Wash solution for hours to clean out everything.

Yes, assuming that most of the barium is "stuck" in the tubing, running nitric or Tuning through all the tubing and having it drip into a tube will clean it....no need for plasma to be on. However, as you know, this would not address any Ba stuck to the spray chamber, injector, or cones. I'd probably worry most about the Injector, as that's the part that I see get the most dirty from samples; Helios spray chambers don't seem to get so dirty as the old v1/v2 glass ones.


Regarding the gut samples: honestly, this is something that your lab will have to decide policy on. On rare occasions, the HIMC has refused to run samples that are too highly contaminated, on the basis that it damages the instrument (ages the detector, etc) and affects other people's runs. At the *very* least, such samples have to be run the day before you plan to fully clean the instrument, so that you're not having to do a full clean every day.

That means that you have to get your customers/researchers to tell you what the heck they did with the samples, including all the way back to where they were coming out of the body. For example, if they did an upper GI imaging series and had someone drink Barium for Xray contrast before sample biopsy, I don't know that such a sample could *ever* be run on a CyTOF. I have no idea what the timeline/kinetics of barium excretion are from the gut: Ba salts are generally insoluble in water....whether a next day or next week sample would be clean enough, I don't know.


Mike
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anitamkant

Master

Posts: 51

Joined: Mon Nov 18, 2013 6:30 am

Post Mon Jul 23, 2018 9:18 pm

Re: Barium contamination cleanup

"Hi James,
Thank you for bringing up an important topic on the forum.
One can run 2% v/v nitric acid on Helios with Plasma On to flush out Barium contamination. Note that we advise that you do not to monitor ion signal during the initial period of nitric acid introduction as the steep rise in the signal from the flushed out Barium may overwhelm the detector.
It is also advisable to run deionized water for 15 minutes thereafter to equilibrate pH of the system.
Regards,
Team Fluidigm"
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JamesW

Contributor

Posts: 37

Joined: Tue Nov 21, 2017 6:59 am

Post Tue Jul 24, 2018 1:58 am

Re: Barium contamination cleanup

Dear All,

Thank you very much for your time and advice. I have Optima Grade Nitric Acid so will add a dilution of that to our cleaning routines. We have asked our users to find out what is happening with their samples but they say that there is no indication that they have had recent barium imaging so it may be quite persistent or coming from an unknown source. We are currently working out a threshold for how contaminated a sample can be before we ask that it not be run.

Best,
James
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mleipold

Guru

Posts: 5792

Joined: Fri Nov 01, 2013 5:30 pm

Location: Stanford HIMC, CA, USA

Post Tue Jul 24, 2018 2:48 pm

Re: Barium contamination cleanup

Hi James,

Are they preparing/staining their own samples? If so, you may want to check their buffers: as has been said many times over the years, buffers are the main source of contamination, particularly barium. You can usually just take an aliquot of the buffer, dilute it 1e3-1e5 times in MilliQ, and run in Liquid mode.

Even if someone uses MilliQ water and metal-free buffer stock like the Rockland PBS, they may still be making them in bottles that have gone through departmental dishwash, which would irreversibly contaminate the bottles with Barium from the commercial soap. Similarly, while I know that a lot of people use regular Gibco PBS for their washes, I've seen lot-to-lot variation in the contamination levels in that buffer.

In some cases, it can come from sample isolation upstream of the customer: I remember one time where a lab that was looking at BAL samples had terrible contamination from the clinical group that was actually doing the lavage and then giving the samples to that second lab. The only way they eventually solved it was by making a bunch of buffer and giving it to the clinical lab for use in the sample preparation.


Mike
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JamesW

Contributor

Posts: 37

Joined: Tue Nov 21, 2017 6:59 am

Post Thu Jul 26, 2018 1:42 am

Re: Barium contamination cleanup

HI Mike,

Thanks for your input. Yes, they prepare all of the samples themselves. I think it may be more intrinsic to the individual samples as in each case they run a batch of samples that they have processed together using the same buffers and only one of them is contaminated. Having said that, I may ask them to check the buffers just in case and also try to find out if there are any sample preparation issues relating to the initial isolation of the tissue samples.

Best,
James

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