The evolution of spectral flow cytometry.

This special issue of Cytometry marks the transition of spectral flow cytometry from an emerging technology into a transformative force that will shape the fields of cytometry and single-cell analysis for some time to come. Tracing its roots to the earliest years of flow cytometry, spectral flow cytometry has evolved from the domain of individual researchers pushing the limits of hardware, reagents, and software to the mainstream, where it is being harnessed and adapted to meet the analytical challenges presented by modern biomedical research. In particular, the current form of spectral flow technology has arisen to address the needs of multiparameter immunophenotyping of immune cells in basic and translational research, and much of the current instrumentation and software reflects the needs of those applications. Yet, the possibilities enabled by high-resolution analysis of the spectral properties of optical absorbance, scatter, and emission have only begun to be exploited. In this brief review, the author highlights the origins and early milestones of single-cell spectral analysis, assesses the current state of instrumentation and software, and speculates as to future directions of spectral flow cytometry technology and applications.