Session: Session 3
The promise of the Orbitrap ASTRAL mass-spectrometer for proteomics analysis
The introduction of the Orbitrap asymmetric track lossless (ASTRAL) analyzer is a major technological leap for MS-based analysis with a scan speed of up to 200 Hz, combining sensitivity, reproducibility, robustness and high throughput.
We performed several experiments to evaluate the performance of the system for bulk and single cell protein identification.
The results were compared with those of the Exploris 480 (E480). Both mass spectrometers were coupled to the Vanquish Neo using an EasySpray source. Data analysis was performed using Proteome Discoverer 3.1.
After injecting 250 ng of HeLa digest, the ASTRAL identified 10,000 proteins in only 30 minutes, while the E480 identified only 7,000 proteins in 60 minutes. We then analyzed different mixtures of 3 commercial digests (HeLa, yeast and E. coli) to measure the accuracy of quantification with fixed and variable fractions for HeLa and the other two species, respectively. We used a 9-minute gradient, corresponding to approximately 160 samples analyzed per day, to monitor ASTRAL's performance in a high-throughput analysis. The three proteomes were identified each time in the different mixtures, with a total of 10,000 proteins identified per mixture. The theoretical ratios were also maintained with a minimum accuracy of 80%. We tested the ASTRAL coupled to Vanquish Neo in single cell conditions against E480 equipped with Field Asymmetric Ion Mobility Spectrometry (FAIMS) and Loload column. 250 pg of the previous digestion mixtures were injected on the ASTRAL, allowing the identification of 3,500 proteins despite the absence of FAIMS.
In conclusion, the ASTRAL surpassed the performance of the E480 in terms of depth, protein identification and proteome coverage, while requiring less material and shorter turnaround time. We believe that the ASTRAL, coupled with the FAIMS, will help to discover new proteins of interest that have never been seen before and to implement proteomics into clinical routine.
We performed several experiments to evaluate the performance of the system for bulk and single cell protein identification.
The results were compared with those of the Exploris 480 (E480). Both mass spectrometers were coupled to the Vanquish Neo using an EasySpray source. Data analysis was performed using Proteome Discoverer 3.1.
After injecting 250 ng of HeLa digest, the ASTRAL identified 10,000 proteins in only 30 minutes, while the E480 identified only 7,000 proteins in 60 minutes. We then analyzed different mixtures of 3 commercial digests (HeLa, yeast and E. coli) to measure the accuracy of quantification with fixed and variable fractions for HeLa and the other two species, respectively. We used a 9-minute gradient, corresponding to approximately 160 samples analyzed per day, to monitor ASTRAL's performance in a high-throughput analysis. The three proteomes were identified each time in the different mixtures, with a total of 10,000 proteins identified per mixture. The theoretical ratios were also maintained with a minimum accuracy of 80%. We tested the ASTRAL coupled to Vanquish Neo in single cell conditions against E480 equipped with Field Asymmetric Ion Mobility Spectrometry (FAIMS) and Loload column. 250 pg of the previous digestion mixtures were injected on the ASTRAL, allowing the identification of 3,500 proteins despite the absence of FAIMS.
In conclusion, the ASTRAL surpassed the performance of the E480 in terms of depth, protein identification and proteome coverage, while requiring less material and shorter turnaround time. We believe that the ASTRAL, coupled with the FAIMS, will help to discover new proteins of interest that have never been seen before and to implement proteomics into clinical routine.