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Flow Cytometry

Flow Cytometry

The development of flow cytometry techniques began in 1934 with initial attempts by Andrew Moldavan, a bacteriologist who counted the cells in a flowing capillary system. Moldavan pumped cells through a capillary tube focused under a microscope for visualization. The invention of photometry allowed the measurement of differentially-stained cells, and Louis Kamentsky developed the first multiparameter flow cytometer in 1965 to identify and count the cancer cells in cervical tissue specimens.
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Updated: Feb 20, 2026

Multicolor Flow Cytometry-based Quantification of Mitochondria and Lysosomes in T Cells

Multicolor Flow Cytometry-based Quantification of Mitochondria and Lysosomes in T Cells

Published on: January 9, 2019

Quantitative Fluorescence Measurements with Multicolor Flow Cytometry.

Lili Wang¹, Adolfas K Gaigalas², James Wood³

¹National Institute of Standards and Technology (NIST), 100 Bureau Drive, Stop 8312, Gaithersburg, MD, 20899, USA. lili.wang@nist.gov.

Methods in Molecular Biology (Clifton, N.J.)

|October 27, 2017

Summary

This summary is machine-generated.

This study presents a new method for quantifying protein biomarkers using multicolor flow cytometry. The procedure ensures accurate, instrument-independent measurements in antibodies bound per cell (ABC).

Keywords:

Antibodies bound per cell CD4+ lymphocytes Compensation Equivalent number of reference fluorophores (ERF)Fluorescence calibration Instrument quality control Instrument sensitivity Multicolor flow cytometry Pulsed light-emitting diode (LED) light source

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Area of Science:

Biotechnology
Clinical Laboratory Science
Immunology

Background:

Multicolor flow cytometry is vital for clinical diagnostics, disease monitoring, and prognostication.
Current quantitative measurement methods lack instrument independence and satisfactory accuracy.
Standardized quantification is needed for reliable biomarker analysis.

Purpose of the Study:

To detail a procedure for quantifying surface and intracellular protein biomarkers using multicolor flow cytometry.
To establish instrument-independent quantitative measurements in antibodies bound per cell (ABC).
To improve the reliability and reproducibility of flow cytometry data in clinical settings.

Main Methods:

Quality control and performance characterization of the multicolor flow cytometer.
Fluorescence calibration using microspheres in equivalent number of reference fluorophores (ERF).
Correction for fluorescence spillover (compensation) and application of a biological reference standard.

Main Results:

A detailed procedure for quantifying protein biomarkers in units of antibodies bound per cell (ABC).
The method enables fluorescence calibration and accurate compensation for reliable measurements.
Successful translation from the ERF scale to the ABC scale using a biological reference standard.

Conclusions:

The described procedure offers a robust method for quantitative biomarker analysis via multicolor flow cytometry.
This approach aims to achieve instrument-independent quantification, overcoming limitations of existing methods.
Ongoing efforts focus on platform-independent biomarker quantification for broader clinical application.