This study introduces a novel mass spectrometry (MS) strategy called narrow-window data-independent acquisition (nDIA), which combines high-resolution MS1 scans with ultra-fast MS/MS scans at ~200 Hz using 2-Th isolation windows. This approach enables comprehensive proteome coverage with high sensitivity and accuracy, surpassing current state-of-the-art MS methods. The nDIA strategy allows for the profiling of over 100 full yeast proteomes per day or 48 human proteomes per day, with deep coverage of ~10,000 human protein groups in half an hour or ~7,000 proteins in 5 minutes. Multi-shot acquisition of offline fractionated samples provides comprehensive coverage of human proteomes in ~3 hours. The method demonstrates high quantitative precision and accuracy, quantifying over 14,000 protein groups in a single half-hour run. The nDIA strategy uses a quadrupole Orbitrap mass spectrometer paired with the asymmetric track lossless (Astral) analyzer, which provides high resolution, sensitivity, and mass accuracy. This combination enables the use of narrow 2-Th DDA-like isolation windows for nDIA, allowing for comprehensive peptide precursor coverage. The method is ideal for high-throughput proteomics and enables the identification of over 170,000 peptide precursors and ~10,000 protein groups in human proteome profiling. The nDIA approach also provides high reproducibility, with a coefficient of variation (CV) below 20% for 90% of precursors and 95% of proteins. The study demonstrates that nDIA enables precise and accurate label-free quantification, with the ability to identify over 2× more proteins and 3.5× more peptide precursors in a 28-min LC gradient compared to the Orbitrap Exploris 480 mass spectrometer. The method also allows for the identification of over 3× more peptides and 2× more protein groups in spectral library-free mode compared to other MS platforms. The nDIA strategy is particularly well-suited for single-cell proteomics research, enabling the identification of ~3,000 proteins from 50 pg and >4,500 proteins from single HeLa cells. The study also highlights the potential of nDIA for systems biology and clinical proteomics, with the ability to analyze large clinical cohorts and conduct biomarker discovery experiments. The method enables the analysis of a yeast strain library consisting of 104 gene knockouts targeting cell cycle, proteasome, DNA damage response, and kinase genes, with consistent quantification of ~4,500 proteins across all biological conditions and replicates. The nDIA approach also enables the comprehensive characterization of the MSA proteome, with the ability to identify over 9,236 protein groups with a median ofThis study introduces a novel mass spectrometry (MS) strategy called narrow-window data-independent acquisition (nDIA), which combines high-resolution MS1 scans with ultra-fast MS/MS scans at ~200 Hz using 2-Th isolation windows. This approach enables comprehensive proteome coverage with high sensitivity and accuracy, surpassing current state-of-the-art MS methods. The nDIA strategy allows for the profiling of over 100 full yeast proteomes per day or 48 human proteomes per day, with deep coverage of ~10,000 human protein groups in half an hour or ~7,000 proteins in 5 minutes. Multi-shot acquisition of offline fractionated samples provides comprehensive coverage of human proteomes in ~3 hours. The method demonstrates high quantitative precision and accuracy, quantifying over 14,000 protein groups in a single half-hour run. The nDIA strategy uses a quadrupole Orbitrap mass spectrometer paired with the asymmetric track lossless (Astral) analyzer, which provides high resolution, sensitivity, and mass accuracy. This combination enables the use of narrow 2-Th DDA-like isolation windows for nDIA, allowing for comprehensive peptide precursor coverage. The method is ideal for high-throughput proteomics and enables the identification of over 170,000 peptide precursors and ~10,000 protein groups in human proteome profiling. The nDIA approach also provides high reproducibility, with a coefficient of variation (CV) below 20% for 90% of precursors and 95% of proteins. The study demonstrates that nDIA enables precise and accurate label-free quantification, with the ability to identify over 2× more proteins and 3.5× more peptide precursors in a 28-min LC gradient compared to the Orbitrap Exploris 480 mass spectrometer. The method also allows for the identification of over 3× more peptides and 2× more protein groups in spectral library-free mode compared to other MS platforms. The nDIA strategy is particularly well-suited for single-cell proteomics research, enabling the identification of ~3,000 proteins from 50 pg and >4,500 proteins from single HeLa cells. The study also highlights the potential of nDIA for systems biology and clinical proteomics, with the ability to analyze large clinical cohorts and conduct biomarker discovery experiments. The method enables the analysis of a yeast strain library consisting of 104 gene knockouts targeting cell cycle, proteasome, DNA damage response, and kinase genes, with consistent quantification of ~4,500 proteins across all biological conditions and replicates. The nDIA approach also enables the comprehensive characterization of the MSA proteome, with the ability to identify over 9,236 protein groups with a median of