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Flow Cytometry01:23

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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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Identifying Microglia and Peripheral Infiltrating Macrophages in the Injured Spinal Cords Using Flow Cytometry
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A two-dimensional suspension array system by coupling field flow fractionation to flow cytometry.

Jishan Li1, Wenwan Zhong

  • 1Department of Chemistry, University of California, Riverside, CA 92521, USA.

Journal of Chromatography. A
|February 5, 2008
PubMed
Summary
This summary is machine-generated.

Flow field flow fractionation (Fl-FFF) coupled with flow cytometry enhances suspension array performance. This size-based separation method is compatible with immunoassay conditions and increases sample throughput.

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Area of Science:

  • Biotechnology
  • Analytical Chemistry
  • Biochemistry

Background:

  • Suspension arrays are widely used for multiplexed immunoassays.
  • Current limitations include sample throughput and assay background.
  • Flow cytometry is a common detection method for suspension arrays.

Purpose of the Study:

  • To improve the performance of suspension arrays by coupling Flow field flow fractionation (Fl-FFF) with flow cytometry.
  • To assess the compatibility of Fl-FFF with immunoassay conditions.
  • To evaluate the impact of Fl-FFF on sample throughput and assay background.

Main Methods:

  • Coupling of Flow field flow fractionation (Fl-FFF) to flow cytometry.
  • Size-based separation of protein-conjugated microspheres using Fl-FFF.
  • Analysis of microspheres using fluorescence measurements and SDS-PAGE.
  • Evaluation of carrier fluid condition tolerance (pH, salt, buffer composition).

Main Results:

  • Fl-FFF successfully separated microspheres based on size.
  • The separation process tolerated a wide range of buffer conditions suitable for immunoassays.
  • Immuno-complex integrity was maintained throughout Fl-FFF separation.
  • Fl-FFF increased sample throughput several-fold by enabling separation of particles with different sizes.
  • Continuous washing in Fl-FFF potentially reduced assay background.

Conclusions:

  • Fl-FFF is a valuable technique for enhancing suspension array performance.
  • The method offers improved sample throughput and reduced assay background.
  • Fl-FFF is compatible with the stringent conditions required for immunoassays.