A major limitation to applying quantitative LC-MS/MS proteomics to small samples, such as single cells, are the losses incured during sample cleanup. To relieve this limitation, we developed a Minimal ProteOmic sample Preparation (mPOP) method for culture-grown mammalian cells. mPOP obviates cleanup and thus eliminates cleanup-related losses while expediting sample preparation and simplifying its automation. Bulk SILAC samples processed by mPOP or by conventional urea-based methods indicated that mPOP results in complete cell lysis and accurate relative quantification. We integrated mPOP lysis with the Single Cell ProtEomics by Mass Spectrometry (SCoPE-MS) sample preparation, and benchmarked the quantification of such samples on a Q-exactive instrument. The results demonstrate low noise and high technical reproducibility. Then, we FACS sorted single U-937, HEK-293, and mouse ES cells into 96-well plates and analyzed them by automated mPOP and SCoPE-MS. The quantified proteins enabled separating the single cells by cell-type and cell-division-cycle phase.
slavovLabI have always hated the career building aspects of pursuing an academic career, and yet I do not have good ideas how to sidestep them.
These aspects aren't unique to academia (seem worse in politics) but are particularly incongruous with what inspired many of us to do research.
slavovLab@skryazhi@NSF I strongly dislike invoking "competition with China" as a motivation -- this is misguided on many levels.
It's hard to predict that this will evolve, and I do not know enough to make an informed prediction. It sounds more promising than menacing.