Abstract
INTRODUCTION: With their noninvasive capabilities for molecular-level analysis of biological tissues and cells, Raman spectroscopy and microscopy have become important tools in medicine and medical diagnostics. METHODS: In this work, we describe a reliable and repeatable technique for Raman spectroscopic profiling of human neutrophils in peripheral blood smears. 1200 single-cell spectra from 12 healthy donors were obtained using Raman microscopy, and the results showed consistent spectral features of key biomolecules such as proteins, nucleic acids, carotenoids, and cytochrome c. RESULTS: We used t-distributed stochastic neighbor embedding and principal component analysis to assess interindividual variability; both techniques showed minimal spectral divergence between donors and high reproducibility. This study lays the methodological groundwork for upcoming Raman microscopy applications enabling non-invasive assessment of immune cell states in health and disease. CONCLUSION: Our results demonstrate the potential of Raman microscopy in clinical diagnostics, specifically for tracking the metabolic states of cells.