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Flow Cytometry Basics

Flow Cytometry: A Basic Introduction. Excellent resource accessible to all levels of experience. Latter chapters are specific to experiment types (DNA content, apoptosis, etc) with earlier chapters providing an excellent basis for newcomers to the field. Data figures include original data links if you have access to FCS express, making the learning exercises interactive. There are also very good references, if a bit dated.

Practical Flow Cytometry: A great guide that covers everything including how a cytometer works, historical context, light interacting with matter and what fluorophores work for a gamut of applications. A bit heavy on physics in places, but that won’t keep you from gleaning critical cytometry knowledge if you can’t digest those parts. It is very compartmentalized, humorous and most chapters are fairly standalone for those with some knowledge. A free PDF version is available online at Beckman Coulter (you need to provide contact information).

General flow cytometry lectures: dated but still very useful from a household name in flow cytometry- J. Paul Robinson and a great starting point for those getting into cytometry that want to know how and why everything works.

General Information and Experiment Design

BD protocols: webinars, tools, technical application notes that can be very helpful.

Beckman Coulter Flow Cytometry Webinars: on-demand webinars that can be very helpful.

Purdue University Cytometry Laboratories: The most extensive listserv for flow cytometry- cytometrists of all levels of experience can ask questions from troubleshooting complex experiments to gathering information regarding cytometer purchasing decisions or DIY fixes. Just remember to search the archives before asking a question that may have already been answered.

Linscott Directory: a very useful directory of antibodies and helpful resource when working on experiment/panel design. Can select many helpful fields to narrow your search (species, fluor, etc.).

UCLA website: with some helpful protocols (general staining, blocking and FcR blocking steps, example protocols, etc.). Several of the protocols are geared towards blood cell preps or are a bit dated but still useful starting points.

Controls and determination of positivity: a helpful paper for people getting started in cytometry
Maecker, HT., and Trotter J. Flow Cytometry Controls, Instrument Setup, and the Determination of Positivity. Cytometry Part A. 2006. 69A:1037–1042.


Microparticles (exosomes, microvessicles, etc.) is a hot topic and there is some lively discussion on how to appropriately measure them. This debate is ongoing and protocol specifics are crucial in achieving reproducible results, with a “consensus” protocol remaining a controversial subject. Below are several references to key papers helping you to get oriented with the ins and outs of flow cytometric measurement of microparticles.

Lacroix et al, High-Sensitivity Flow Cytometry Provides Access to Standardized Measurement of Small-Size Microparticles, Arterioscler Thromb Vasc Biol. 2012 Apr;32(4):1054-8.

Larson, M.C., M.R. Luthi, N. Hogg and C.A. Hillery. 2013 Calcium-Phosphate microprecipitates mimic microparticles when examined with flow cytometry. Cytometry Part A. 83A: 242-250.

Nolan, J.P., and S.A. Stoner. 2013 A trigger channel threshold artifact in nanoparticle analysis. Cytometry Part A. 83A: 301-305.

Van der Pol et al., Single vs. swarm detection of microparticles and exosomes by flow cytometry. J Thromb Haemost, 2012. 10(5):919-30

Lee, R.D., et al., Pre-analytical and analytical variables affecting the measurement of plasma-derived microparticle tissue factor activity. Thromb Res, 2011.

Ayers, L., et al., Measurement of circulating cell-derived microparticles by flow cytometry: sources of variability within the assay.Thromb Res, 2011. 127(4): p. 370-7.

Yuana, Y., R.M. Bertina, and S. Osanto, Pre-analytical and analytical issues in the analysis of blood microparticles. Thromb Haemost, 2011. 105: p. 396-408.

Lacroix R, Robert S, Poncelet P, Dignat-George F. Overcoming limitations of microparticle measurement by flow cytometry. Semin Thromb Hemost, 2010. 36(8):807-18.

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