Ficoll-iodine levels
Hi all,
I finally got around to testing aliquots of Ficoll and Lymphoprep solutions by Liquid mode Helios CyTOF. This was to confirm the article referenced in Gaudilliere et al-2015 that Ficoll has large amounts of an iodine-containing preservative.
Ficoll TDS:
https://www.gelifesciences.com/gehcls_i ... 233425.pdf
Lymphoprep MSDS:
http://www.stemcell.com/~/media/Technic ... .pdf?la=en
https://en.wikipedia.org/wiki/Diatrizoic_acid
Someone asked me if I had tested the Lymphoprep solution from Stem Cell as an alternative. So, I bought some of that and tested it too.
I made a dilution series of the Lymphoprep or Ficoll in MilliQ water, and ran in Liquid Mode on a Helios. I monitored 89-209 (except 103, by mistake).
The bottom line: both solutions contain sodium diatrizoate, which contains 3 iodine atoms per molecule. At 1e6 (1 million-fold!) dilution, the Iodine signal was around 2.5 *million* Dual units. For comparison, this is roughly the same as the Tb in the Tuning solution on that Helios instrument.
I'm sure that skill at removing the PBMC layer while not taking any Ficoll would make a difference in how much iodine reagent comes along with the cells, and I assume this would be further reduced by washing steps after. Alternatively, StemCell's SepMate tubes may help: while they still use Ficoll or Lymphoprep, the PBMCs wind up above a separation membrane from the Ficoll and minimizes the likelihood of getting Ficoll in your PBMCs.
But the bottom line is, I'm not sure what this means in terms of consistency of iodine contamination background levels in PBMC assays......particularly if you're wanting to use IdU as a cell cycle reagent.
I don't use IdU in my assays: maybe someone with experience using it can comment on what their backgrounds are like (both magnitude and variability) from PBMCs?
Mike
I finally got around to testing aliquots of Ficoll and Lymphoprep solutions by Liquid mode Helios CyTOF. This was to confirm the article referenced in Gaudilliere et al-2015 that Ficoll has large amounts of an iodine-containing preservative.
Ficoll TDS:
https://www.gelifesciences.com/gehcls_i ... 233425.pdf
Lymphoprep MSDS:
http://www.stemcell.com/~/media/Technic ... .pdf?la=en
https://en.wikipedia.org/wiki/Diatrizoic_acid
Someone asked me if I had tested the Lymphoprep solution from Stem Cell as an alternative. So, I bought some of that and tested it too.
I made a dilution series of the Lymphoprep or Ficoll in MilliQ water, and ran in Liquid Mode on a Helios. I monitored 89-209 (except 103, by mistake).
The bottom line: both solutions contain sodium diatrizoate, which contains 3 iodine atoms per molecule. At 1e6 (1 million-fold!) dilution, the Iodine signal was around 2.5 *million* Dual units. For comparison, this is roughly the same as the Tb in the Tuning solution on that Helios instrument.
I'm sure that skill at removing the PBMC layer while not taking any Ficoll would make a difference in how much iodine reagent comes along with the cells, and I assume this would be further reduced by washing steps after. Alternatively, StemCell's SepMate tubes may help: while they still use Ficoll or Lymphoprep, the PBMCs wind up above a separation membrane from the Ficoll and minimizes the likelihood of getting Ficoll in your PBMCs.
But the bottom line is, I'm not sure what this means in terms of consistency of iodine contamination background levels in PBMC assays......particularly if you're wanting to use IdU as a cell cycle reagent.
I don't use IdU in my assays: maybe someone with experience using it can comment on what their backgrounds are like (both magnitude and variability) from PBMCs?
Mike