Wednesday, March 27, 2019
The Merosome Proteome!
Throughout this blog you'll find whining about malaria proteomics. You may see it forever, until we finally beat this ancient disease. Any time I feel like I'm really getting good at this -- or the next generation of instrumentation is finally going to be the thing to blow this field wide open -- I end up deeply humbled by the experience. The people trudging through this full time will always have my respect.
And -- this new study from a lot of people I know -- and a name or two I hope to know soon -- just opened up an aspect of this frustrating organism that I would have thought impossible -- the MEROSOME Proteome!
Stolen image from this amazing text-book level Nature Review from 2008
The merosome is a stage between the Plasmodium leaving the liver (where they hide in low numbers) and work their way into the blood stream while somehow avoiding all immune responses. I don't think the mechanism is well understood at all, but the impression is that they hijack the membranes of liver cells and use those as their escape vessels.
To be clear -- this is a RIDICULOUSLY SMALL number of parasites. Ridiculously small.
How'd they get enough to work with? By doing an absurd amount of work. They grew one of the mouse model species of Plasmodium in cells in culture. By using a combination of microscopy and PCR -- they captured the merosomes and enriched them.
They normalize the samples by the genome amplification numbers. Ever had so little material that you had to AMPLIFY THE DNA to figure out how many cells were there? Me either -- but what a great idea. Honestly -- I'd guess if you had that little material -- even today's best MS technology wouldn't detect a thing -- and I'd be wrong.
This team uses SCX stage tips to fractionate the material -- and used single shot LC-MS (50cm EasySpray on a Lumos) using a method built for sensitivity (30,000 resolution MS/MS + 150ms max fill time -- not sure if max fill time override "use all available fill" was utilized).
Obviously there is going to be a lot going on here -- cell culture contaminants, human cell proteins, etc., so they included all those in a data analysis -- and scored almost 2,000 protein IDs(!!) from a phase of the malaria disease I wouldn't have believed possible to work with if I hadn't read this today.....
The downstream analysis might be equally impressive as the upstream work -- making this just a marvelous and inspiring study.