Overview

2024-05-13

Here, we are evaluating the performance of Astral compared to the Orbitrap for DIA quantitation. To do this, we have two main experiments:

Matrix-matched calibration curve to evaluate quantitative precision and accuracy in a HeLa cell lysate
Plasma proteome measurements to showcase the potential of Astral and a new extracellular vesicle enrichment strategy to reach new depths of coverage in a single shot

mzML files are not included on this page due to space considerations and the fact that they are functionally the same as raw files. mzML files can be generated using the ProteoWizard Docker image available on Docker Hub at “ proteowizard/pwiz-skyline-i-agree-to-the-vendor-licenses:3.0.22335-b595b19”.

To re-generate the files in MSConvert, the following command can be used: msconvert *.raw --simAsSpectra --ignoreUnknownInstrumentError --filter "peakPicking true"

Below is a summary of the run types.

File prefix	Sample	Analyzer	Normalized AGC Target	Maximum injection time	Isolation Window	Mass range	Column	Gradient length (min)
20230403_OLEP08_EV_1ug_MB_30min_AS_10ms_4Th_	EV plasma	Astral	500%	10 ms	4 Th	400-1000	110 cm	30
20230403_OLEP08_EV_1ug_MB_30min_AS_3p5ms_2Th_	EV plasma	Astral	500%	3.5 ms	2 Th	400-1000	110 cm	30
20230403_OLEP08_TP_1ug_MB_30min_AS_10ms_4Th_	Total plasma	Astral	500%	10 ms	4 Th	400-1000	110 cm	30
20230403_OLEP08_TP_1ug_MB_30min_AS_3p5ms_2Th_	Total plasma	Astral	500%	3.5 ms	2 Th	400-1000	110 cm	30
20230404_OLEP08_EV_1ug_MB_60min_AS_15ms_4Th_	EV plasma	Astral	500%	15 ms	4 Th	400-1000	110 cm	60
20230404_OLEP08_EV_1ug_MB_60min_AS_dDIA_15ms_2Th_	EV plasma	Astral	500%	15 ms	2 Th	300 Th dDIA	110 cm	60
20230404_OLEP08_TP_1ug_MB_30min_AS_dDIA_20ms_4Th_	Total plasma	Astral	500%	20 ms	4 Th	300 Th dDIA	110 cm	30
20230404_OLEP08_TP_1ug_MB_60min_AS_15ms_4Th_	Total plasma	Astral	500%	15 ms	4 Th	400-1000	110 cm	60
20230404_OLEP08_TP_1ug_MB_60min_AS_dDIA_15ms_2Th_	Total plasma	Astral	500%	15 ms	2 Th	300 Th dDIA	110 cm	60
20230405_OLEP08_EV_1ug_MB_30min_AS_dDIA_20ms_4Th_	EV plasma	Astral	500%	20 ms	4 Th	300 Th dDIA	110 cm	30
20230405_OLEP08_EV_1ug_MB_60min_AS_23ms_2Th_ChrLib_	EV plasma	Astral	500%	23 ms	2 Th	100 Th GPF	110 cm	60
20230405_OLEP08_EV_1ug_MB_60min_OT_23ms_2Th_ChrLib_	EV plasma	Orbitrap	800%	23 ms	2 Th	100 Th GPF	110 cm	60
20230406_OLEP08_MMCC_1ug_MB_24min_AS_10ms_4Th_	MMCC	Astral	500%	10 ms	4 Th	400-1000	110 cm	24
20230406_OLEP08_MMCC_1ug_MB_24min_AS_3p5ms_2Th_	MMCC	Astral	500%	3.5 ms	2 Th	400-1000	110 cm	24
20230406_OLEP08_MMCC_1ug_MB_24min_AS_dDIA_10ms_2Th_	MMCC	Astral	500%	10 ms	2 Th	300 Th dDIA	110 cm	24
20230406_OLEP08_MMCC_1ug_MB_24min_OT_75Th_23ms_2Th_	MMCC	Orbitrap	800%	23 ms	2 Th	500-575	110 cm	24
20230410_OLEP08_HeLa_1ug_MB_24min_AS_23ms_2Th_ChrLib_	200 ng HeLa	Astral	500%	23 ms	2 Th	100 Th GPF	110 cm	24

MMCC: Matrix-matched calibration curve of HeLa into SILAC labeled HeLa. Consists of 9 dilution points injected in triplicate.

EV plasma: 1 ug EV enriched plasma

Total plasma: 1 ug of total plasma

GPF: 6 gas phase fraction runs, each covering 100 Da range

dDIA: dynamic DIA