Dataset Information

Validation of Thin-diaPASEF for maximizing proteome coverage

ABSTRACT: We optimized the parameters of CCS range, polygon scan region, ramp time, and isolation window width to achieve high-depth proteome analysis using the timsTOF HT. As a result, we developed Thin-diaPASEF, which was optimized with a CCS range of 0.7–1.3, a ramp time of 150 ms, an isolation window width of 26 Th (with an overlap width of 1 Th), and a polygon area focused on regions where precursors accumulate. Thin-diaPASEF was then compared with existing methods, including py-diAID PASEF, slice-PASEF, and synchro-PASEF. The results demonstrated that Thin-diaPASEF achieved superior protein identification. Performance evaluation of Thin-diaPASEF revealed the identification of approximately 9,400 proteins with a sample injection of 500 ng and an analysis time of 100 min. Finally, we applied Thin-diaPASEF to plasma proteome analysis. We compared three sample preparation methods: an improved LEL method based on our previously reported plasma and serum preparation approach, Seer’s nanoparticle-based method, and the conventional Top14 column-based method. As a result, the improved LEL method successfully identified 5,378 proteins at an analysis throughput of 24 samples per day.

ORGANISM(S): Homo Sapiens (human) Cellular Organisms

SUBMITTER: Yusuke Kawashima

PROVIDER: PXD061613 | JPOST Repository | Sun Aug 10 00:00:00 BST 2025

REPOSITORIES: jPOST

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Dataset's files

Source:

			Action	DRS
	HEK100ng_100ms_07-13_default-diaPASEF-1.d.zip	Other
	HEK100ng_100ms_07-13_default-diaPASEF-2.d.zip	Other
	HEK100ng_100ms_07-13_default-diaPASEF-3.d.zip	Other
	HEK100ng_100ms_07-13_default-diaPASEF-4.d.zip	Other
	HEK100ng_100ms_07-13_default-diaPASEF-5.d.zip	Other

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Publications

Thin-diaPASEF: diaPASEF for maximizing proteome coverage in single-shot proteomics.

Konno Ryo R Ishikawa Masaki M Nakajima Daisuke D Inukai Kaori K Ohara Osamu O Kawashima Yusuke Y

DNA research : an international journal for rapid publication of reports on genes and genomes 20250701 4

Proteomics using mass spectrometry (MS) has significantly advanced, offering deep insights into complex proteomes. The timsTOF MS platform with its parallel accumulation-serial fragmentation (PASEF) technology has achieved high scan speeds and high-quality spectra. Bruker's timsTOF HT, which features TIMS-XR technology, offers an improved dynamic range and analysis depth, supporting high sample loadings. Moreover, various improvements to the data-independent acquisition method based on the PASEF ...[more]

PMID: 40704459

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Project description:Background: Circular RNA (circRNA) has been reported to be involved in the pathogenesis of cardiovascular disease (CVD), however, it is unclear whether circRNA carried by exosomes (exos) can be used as biomarkers for chronic coronary syndrome (CCS). This study aimed to investigate the expression pattern of exosomal circRNAs in the plasma of CCS patients, and to screen exo-circRNAs that can act as biomarkers for the diagnosis of CCS. Methods: High-throughput sequencing technology was carried out in the plasma exosomal RNA of 15 CCS patients and 15 non-cardiac chest pain patients (NCCP, control group) (sequencing cohort) to screen for differentially expressed circRNAs. Selected differentially expressed exo-circRNAs were further verified by real time polymerase chain reaction (RT-PCR) in a small-sample cohort (derivation cohort) and a large-sample cohort (validation cohort). Receiver operating characteristic (ROC) curve analysis was performed to evaluate the diagnostic value of exo-circRNAs for CCS patients. Results: By high-throughput sequencing of circRNAs in 15 CCS patients and 15 NCCP patients, 276 circRNAs differentially expressed in plasma exosomes of CCS patients were screened by using |log2(Fold change) |>1 and q-value<0.001, and 103 were up-regulated and 173 were down-regulated. Among the 103 up-regulated circRNAs, 5 circRNAs with high expression levels and significantly increased in plasma exosomes of CCS patients were selected for validation. Twice RT-PCR validation demonstrated that exo-hsa_circ_0075269 and exo-hsa_circ_0000284 were significantly up-regulated in patients with CCS (P<0.001). Circulating exo-hsa_circ_0075269 and exo-hsa_circ_0000284 yielded the area under the curve (AUC) values of 0.761 (P<0.001, 95%CI=0.669, 0.852) and 0.623 (P=0.015, 95%CI=0.522, 0.724) for CCS, respectively by ROC curve analysis. Moreover, AUC of hsa_circ_0075269 combined with hsa_circ_0000284 for diagnosing CCS was 0.741 (P<0.001, 95%CI=0.667, 0.850). Conclusion: The expression profile of circRNA in plasma exosomes of patients with CCS was significantly different from that of the control group. Plasma exo-hsa_circ_0075269 and exo-hsa_circ_0000284 have the potential to be new biomarkers for CCS diagnosis.