Biology subjects

Batruch, I.

Publications and source records attributed to Batruch, I..

6 recordsLinked to original sources

bioRxiv2026

Scanning DIA on the ZenoTOF 8600 system enables ultra-sensitive and quantitative proteomics from single cells to post-translational modifications in a compact platform

Mass spectrometry-based proteomics increasingly demands platforms that combine quantitative rigor with the discovery capabilities of accurate mass systems. Here we present the ZenoTOF 8600 system, a compact mass spectrometry system that integrates enhanced ion capture and transmission optics with an optical detection system, Zeno trap-enhanced MS/MS, electron-activated dissociation, and scanning quadrupole data-independent acquisition (ZT Scan DIA). We show that ZT Scan DIA outperforms conventional variable-window DIA (Zeno SWATH DIA) in both identifications and quantitative reproducibility, and demonstrate the platforms versatility across proteomics applications: thousands of protein groups from bulk samples at up to 500 samples per day, single-cell proteomics yielding up to 4,700 proteins, accurate ratio recovery in mixed-species quantitative benchmarks, low-attomole targeted quantitation, and detection of disease-relevant phosphorylation in a Parkinsons disease cellular model using complementary CID and EAD fragmentation. The instruments compact footprint makes it attractive for settings where both analytical breadth and operational robustness are required.

biophysics↗

bioRxiv2026

LFQ Benchmark Dataset - Generation Beta: Assessing Modern Proteomics Instruments and Acquisition Workflows with High-Throughput LC Gradients

Recent advances in liquid chromatography-mass spectrometry (LC-MS) have accelerated the adoption of high-throughput workflows that deliver deep proteome coverage using minimal sample amounts. This trend is largely driven by clinical and single-cell proteomics, where sensitivity and reproducibility are essential. Here, we extend our previous benchmark dataset (PXD028735) using next-generation LC-MS platforms optimized for rapid proteome analysis. We generated an extensive DDA/DIA dataset using a human-yeast-E. coli hybrid proteome. The proteome sample was distributed across multiple laboratories together with standardized analytical protocols specifying two short LC gradients (5 and 15 min) and low sample input amounts. This dataset includes data acquired on four different platforms, and features new scanning quadrupole-based implementations, extending coverage across different instruments and acquisition strategies. Our comprehensive evaluation highlights how technological advances and reduced LC gradients may affect proteome depth, quantitative precision, and cross-instrument consistency. The release of this benchmark dataset via ProteomeXchange (PXD070049 and PXD071205), allows for the acceleration of cross-platform algorithm development, enhance data mining strategies, and supports standardization of short-gradient, high-throughput LC-MS-based proteomics.

bioinformatics↗

bioRxiv2025

Performance Characteristics of Zeno Trap Scanning DIA for Sensitive and Quantitative Proteomics at High Throughput

Proteomic experiments, particularly those addressing dynamic proteome properties, time series, or genetic diversity, require the analysis of large sample numbers. Despite significant advancements in proteomic technologies in recent years, further improvements are needed to accelerate measurement and enhance proteome coverage and quantitative performance. Previously, we demonstrated that incorporating a scanning MS2 dimension into data-independent acquisition methods (Scanning SWATH, or more generally scanning DIA) but also ion trapping, improves analytical depth and quantitative performance, especially in proteomic methods using fast chromatography. Here, we evaluate the scanning DIA approach combined with ion trapping via the Zeno trap in a method termed ZT Scan DIA, using a prototype Zeno trap Q-TOF instrument (SCIEX). Applying this method to established proteome standards across various analytical setups, enabling intermediate to high sample throughput, we observed a 30-40% increase in identified precursors. This enhancement extended to overall protein identification and precise quantification. Furthermore, ZT Scan DIA effectively eliminated quantitative bias, as demonstrated by its ability to deconvolute proteomes in multi-species mixtures. We propose that ZT Scan DIA can be used for a broad range of applications in proteomics, particularly in studies requiring high quantitative precision with low sample input and high-throughput workflows. Significance StatementThe advent of faster DIA proteomics methods paved the way for investigating increasingly large sample series, including patient cohorts and strain collections containing thousands of samples. Yet, recent improvements in DIA methods still entailed compromises between analytical sensitivity and selectivity. The presented combination of a scanning quadrupole with fast ion trapping in a Zeno Trap, coined ZT Scan DIA increases quantitative precision and accuracy in fast proteomics experiments. These features of ZT Scan DIA may benefit applications that deal with low input samples and high-throughput proteomic workflows in biomedical cohort studies and systems biology.

systems biology↗

bioRxiv2022

High-throughput proteomics of nanogram-scale samples with Zeno SWATH DIA

The ability to conduct high-quality proteomic experiments in high throughput has opened new avenues in clinical research, drug discovery, and systems biology. Next to an increase in quantitative precision, recent developments in high-throughput proteomics have also gained proteomic depth, to the extent that earlier gaps between classic and high-throughput experiments have significantly narrowed. Here we introduce and benchmark Zeno SWATH, a data-independent acquisition technique that employs a linear ion trap pulsing (Zeno trap pulsing) in order to increase proteomic depth and dynamic range in proteomic experiments. Combined with the high acquisition speed, these gains in sensitivity are particularly attractive for conducting high-throughput proteomics experiments with high chromatographic flow rates and fast gradients. We demonstrate that when combined with either micro-flow- or analytical-flow-rate chromatography, Zeno SWATH increases protein identification in complex samples 5- to 10-fold when compared to current SWATH acquisition methods on the same instrument. Using 20-min micro-flow chromatography, Zeno SWATH identified > 6,000 proteins from a 62.5 ng load of human cell lysate with more than 5,000 proteins consistently quantified in triplicate injections with a median CV of 6%. Using 5-min analytical-flow-rate chromatography (800 {micro}l/min), Zeno SWATH identified 4,907 proteins from a triplicate injection of 2 {micro}g of a human cell lysate; or more than 3,000 proteins from 250 ng tryptic digest. Zeno SWATH hence facilitates precise proteomic experiments with small sample amounts using a fast and robust high flow-rate chromatographic method, broadening the application space that requires precise proteomic experiments on a large scale.

biochemistry↗

bioRxiv2020

Normothermic Ex-vivo Kidney Perfusion in a Porcine Auto-Transplantation Model Preserves the Expression of Key Mitochondrial Proteins: An Unbiased Proteomics Analysis

Normothermic ex-vivo kidney perfusion (NEVKP) results in significantly improved graft function in porcine auto-transplant models of DCD injury compared to static cold storage (SCS); however, the molecular mechanisms underlying these beneficial effects remain unclear. We performed an unbiased proteomics analysis of 28 kidney biopsies obtained at 3 time points from pig kidneys subjected to 30-minutes of warm ischemia, followed by 8 hours of NEVKP or SCS, and auto-transplantation. 70/6593 proteins quantified were differentially expressed between NEVKP and SCS groups (FDR<0.05). Proteins increased in NEVKP mediated key metabolic processes including fatty acid {beta}-oxidation, the TCA-cycle and oxidative phosphorylation. Comparison of our findings with external datasets of ischemia-reperfusion, and other models of kidney injury confirmed that 47 of our proteins represent a common signature of kidney injury reversed or attenuated by NEVKP. We validated key metabolic proteins (ETFB, CPT2) by immunoblotting. Transcription factor databases identified PPARGC1A, PPARA/G/D and RXRA/B as the upstream regulators of our dataset, and we confirmed their increased expression in NEVKP with RT-PCR. The proteome-level changes observed in NEVKP mediate critical metabolic pathways that may explain the improved graft function observed. These effects may be coordinated by PPAR-family transcription factors, and may represent novel therapeutic targets in ischemia-reperfusion injury.

molecular biology↗

bioRxiv2020

Proteomics Reveals Extracellular Matrix Injury in the Glomeruli and Tubulointerstitium of Kidney Allografts with Early Antibody-Mediated Rejection

Antibody-mediated rejection (AMR) accounts for >50% of kidney allograft losses. AMR is caused by donor-specific antibodies (DSA) against HLA and non-HLA antigens in the glomeruli and the tubulointerstitium, which together with inflammatory cytokines such as tumor necrosis factor alpha (TNF) and interferon gamma (IFN{gamma}), trigger graft injury. Unfortunately, the mechanisms governing cell-specific injury in AMR remain unclear. We studied 30 for-cause kidney biopsies with early AMR, acute cellular rejection or acute tubular necrosis ( non-AMR). We laser-captured and microdissected glomeruli and tubulointerstitium, and subjected them to unbiased proteome analysis. 120/2026 glomerular and 180/2399 tubulointerstitial proteins were significantly differentially expressed in AMR vs. non-AMR biopsies (P<0.05). Basement membrane and extracellular matrix (ECM) proteins were significantly decreased in both AMR compartments. We verified decreased glomerular and tubulointerstitial LAMC1 expression, and decreased glomerular NPHS1 and PTPRO expression in AMR. Cathepsin-V (CTSV) was predicted to cleave ECM-proteins in the AMR glomeruli, and CTSL, CTSS and LGMN in the tubulointerstitium. We identified galectin-1, an immunomodulatory protein upregulated in the AMR glomeruli and linked to the ECM. Anti-HLA class-I antibodies significantly increased CTSV expression, and galectin-1 expression and secretion, in human glomerular endothelial cells. Glutathione S-transferase omega-1 (GSTO1), an ECM-modifying enzyme, was significantly increased in the AMR tubulointerstitium, and in TNF-treated proximal tubular epithelial cells. IFN{gamma} and TNF significantly increased CTSS and LGMN expression in these cells. Basement membranes are often remodeled in chronic AMR, and we demonstrated that this remodeling begins early in glomeruli and tubulointerstitium. Targeting ECM-remodeling in AMR may represent a new therapeutic opportunity. SIGNIFICANCE STATEMENTAntibody-mediated rejection (AMR) accounts for >50% of kidney allograft loss, and is caused by donor-specific antibodies against HLA antigens, which induce maladaptive responses in the kidney glomeruli and tubulointerstitium. This is the first unbiased proteomics analysis of laser-captured/microdissected glomeruli and tubulointerstitium from 30 indication kidney biopsies with early AMR, acute cellular rejection or acute tubular necrosis. >2,000 proteins were quantified in each compartment. We discovered that basement membrane and extracellular matrix (ECM) proteins were significantly decreased in both AMR compartments. Two ECM-modifying proteins, LGALS1 and GSTO1, were significantly increased in glomeruli and tubulointerstitium, respectively. LGALS1 and GSTO1 were upregulated by anti-HLA antibodies or AMR-related cytokines in primary kidney cells, and may represent therapeutic targets to ameliorate ECM-remodeling in AMR.

molecular biology↗