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Reg: Maxpar Direct Immune Profile Assay 30 Markers

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komal2000

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Posts: 11

Joined: Mon Aug 29, 2016 12:55 pm

Location: University of Helsinki, Finland

Post Tue Feb 11, 2020 11:24 am

Reg: Maxpar Direct Immune Profile Assay 30 Markers

Dear CyTOF people,

Has any one used Maxpar Direct Immune Profile Assay 30 Markers kit?. Soon planing to stain the cells using this kit but I have a question before staining. The protocol provided by the fluidigm specifically for this kit: after surface staining fixation with 1.6% PFA is included before adding intercalator in Fix/Perm buffer. my question is do I have to fixation step before adding Ir in Fix/Perm buffer? I was thinking adding Fix/Perm buffer is enough to fix the cells both at the surface and intercellular?


Sincerely
Komal
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Verhoeff

Participant

Posts: 16

Joined: Wed Mar 02, 2016 10:25 am

Post Mon Feb 17, 2020 9:31 am

Re: Reg: Maxpar Direct Immune Profile Assay 30 Markers

Hi Komal,

It is strongly recommended to do include the fixation step with fresh 1.6% PFA, we always include it. The fixative agent in the Fix/Perm buffer degrades over time after repeated exposure to oxygen (by opening the bottle). Multiple labs, as I believe discussed here on this forum, noticed poor fixation of antibodies on cells over time. By fixing your antibodies+metal polymers to the cells with fresh PFA, you ensure proper signal.

Practically, we made aliquots of 250uL of our 16% PFA stocks and store them at -20°C.

Cheers,
Jan
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komal2000

Participant

Posts: 11

Joined: Mon Aug 29, 2016 12:55 pm

Location: University of Helsinki, Finland

Post Tue Feb 18, 2020 7:25 am

Re: Reg: Maxpar Direct Immune Profile Assay 30 Markers

Thanks Jan for the information

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