Fountain Flow Cytometry

Paul Johnson1

1 Department of Physics and Astronomy, University of Wyoming, Laramie, Wyoming
Publication Name:  Current Protocols in Cytometry
Unit Number:  Unit 1.26
DOI:  10.1002/0471142956.cy0126s60
Online Posting Date:  April, 2012
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Fountain Flow Cytometry (FFC) is a simple and inexpensive technology that is adaptable to situations requiring detection and enumeration of cells/organisms at low concentrations, but is limited to particles of relatively high fluorescence intensity. This work presents the basic physics behind the novel scheme Fountain Flow Cytometry employs for the detection of target particles, a hybrid of conventional flow cytometry and video epifluorescence microscopy. The method is based on LED‐induced fluorescence of labeled particles and requires no filtration step. Unlike conventional flow cytometry, the resulting fluorescence is measured with a digital camera as the measured sample flows toward the camera along the optical axis. An automated target particle recognition and enumeration computer program, Biocount, is used to count particles. FFC allows for detection of target particles in transparent and translucent fluids, such as environmental water, blood, and beverages. In addition, FFC can be used for detection of target particles in the presence of high photometric background, including unbound fluorescent dye. This facilitates use of the technique in situations where cells are unwashed. Current applications extend, but are not limited to, particles from µm‐size bacteria to multi‐millimeter‐sized multicellular organisms. Curr. Protoc. Cytom. 60:1.26.1‐1.26.14. © 2012 by John Wiley & Sons, Inc.

Keywords: Fountain Flow Cytometry; flow cytometry; Biocount

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Table of Contents

  • Introduction
  • Computer‐Automated Detection and Enumeration
  • The Physics of Fountain Flow Cytometry
  • Optimization of the Signal‐to‐Noise Ratio of Intensity Measurements
  • Volumetric Throughput, Particle Counting Rate, and Coincident Events
  • Sample Dilution Considerations
  • Summary and Outlook
  • Literature Cited
  • Figures
  • Tables
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Literature Cited

Literature Cited
   Depth of field, 2011. (accessed August 9, 2011).
   Fini, J.B., Pallud‐Mothré, S., Le Mével, S., Palmier, K. Havens, C., Garec, E., LeBrun, M., Mataix, V., Demeneix, B.A., Turque, N., and Johnson, P.E. 2009. An innovative continuous flow system for monitoring heavy metal pollution in water using transgenic Xenopus laevis tadpoles. Environ. Sci. Technol. 43:8895‐8900.
   Geiger counter and counting statistics, 1999.∼niel/astro485/derivations/geiger1.pdf (accessed August 8, 2011).
   Johnson, P.E. 2006. Method and system for counting particles in a laminar flow with an imaging device. U.S. Patent Application No. 11/328,033.
   Johnson, P.E. 2007. High resolution imaging fountain flow cytometry, U.S. Patent No. 7,161,665.
   Johnson, P.E, Deromedi, A.J., Lebaron, P., Catala, P., and Cash, J. 2006. Rapid detection and enumeration of Escherichia coli in aqueous samples using Fountain Flow Cytometry. Cytometry A. 69:1212‐1221.
   Johnson, P.E., Deromedi, A.J., Lebaron, P., Catala, P., Havens, C., and Pougnard, C. 2007a. High throughput, real‐time detection of Naegleria lovaniensis in natural river water using LED‐illuminated Fountain Flow Cytometry, J. Appl. Microbiol. 103:700‐710.
   Johnson, P.E., Havens, C.M., and Johnson, J.F. 2007b. Real‐time, low‐cost detection of individual fungal cells in blood using Fountain Flow™ cytometry. Critical Care 11:32.
   Shapiro, H.M. 2003. Practical Flow Cytometry, 4th edition. John Wiley & Sons, Hoboken, N.J.
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