biostudies-literatureMomenzadeh ANIH National Institute of General Medical SciencesNIGMS NIH HHS100973https://www.ebi.ac.uk/biostudies/studies/S-EPMC12534698biostudies-literatureUnknown5(9)Over the past 2 to 3 years, mass-spectrometry-based single-cell proteomics (SCP) has experienced transformative improvements in microfluidic and robotic sample preparation, innovative MS1- and MS2-based multiplexing strategies, and specialized hardware (e.g., timsTOF Ultra 2, Astral), which have dramatically boosted sensitivity, throughput, and proteome coverage from picogram-level protein inputs. Concurrently, tailored computational workflows that encompass normalization, imputation, and no-code platforms have addressed pervasive missing data challenges and standardized analyses, collectively enabling high-throughput, reproducible profiling of cellular heterogeneity. This minireview summarizes the latest progress in SCP technology and software solutions, highlighting how the closer integrCell genomicsSingle-cell proteomics using mass spectrometry.PMC12534698R35GM142502R35 GM142502Momenzadeh AMeyer JGfalseSingle-cell proteomics using mass spectrometry.Over the past 2 to 3 years, mass-spectrometry-based single-cell proteomics (SCP) has experienced transformative improvements in microfluidic and robotic sample preparation, innovative MS1- and MS2-based multiplexing strategies, and specialized hardware (e.g., timsTOF Ultra 2, Astral), which have dramatically boosted sensitivity, throughput, and proteome coverage from picogram-level protein inputs. Concurrently, tailored computational workflows that encompass normalization, imputation, and no-code platforms have addressed pervasive missing data challenges and standardized analyses, collectively enabling high-throughput, reproducible profiling of cellular heterogeneity. This minireview summarizes the latest progress in SCP technology and software solutions, highlighting how the closer integr2025-01-01T00:00:00Z2025 Sep2026-05-09T03:22:35.932Z2026-05-09T03:12:11.327ZS-EPMC125346984084045210.1016/j.xgen.2025.100973