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Related Concept Videos

Flow Cytometry01:23

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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DNA Microarrays02:34

DNA Microarrays

Microarrays are high-throughput and relatively inexpensive assays that can be automated to analyze large quantities of data at a time. They are used in genome-wide studies to compare gene or protein expression under two varied conditions, such as healthy and diseased states. Microarrays consist of glass or silica slides on which probe molecules are covalently attached through surface functionalization. Most commonly, the slides are prepared through the chemisorption of silanes to silica...

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SFMAC: a novel method for analyzing multiple parameters on lymphocytes with a single fluorophore in cell-microarray

Kazuto Tajiri1, Hiroyuki Kishi, Tatsuhiko Ozawa

  • 1Department of Immunology, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama, Toyama, Japan.

Cytometry. Part a : the Journal of the International Society for Analytical Cytology
|November 18, 2008
PubMed
Summary

This study introduces a novel single-cell analysis method using one fluorophore and photobleaching. This technique allows repeated staining and analysis of the same cells, expanding cytometry capabilities.

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Area of Science:

  • Biotechnology
  • Cell Biology
  • Immunology

Background:

  • Multiplex analysis of single cells requires specific fluorophore and optical filter combinations.
  • Increasing demand for multiplex cell analysis drives the need for new fluorophores and techniques.

Purpose of the Study:

  • To develop a novel method for analyzing numerous cells with multiple parameters on a single-cell basis using a single fluorophore.
  • To enable repeated staining and analysis of the same cells with different markers.

Main Methods:

  • Cells were arrayed onto a microwell chip (45,000 microwells) and stained with a phycoerythrin (PE)-conjugated antibody.
  • Single cells were analyzed using a cell-scanner, followed by photobleaching of PE molecules with blue light.
  • Repeated staining and analysis of the same cells were performed using PE after photobleaching.

Main Results:

  • The developed method allows for the analysis of multiple markers on single cells using only one fluorophore.
  • Successful application in analyzing lymphocytes from peripheral blood for cytokine secretion, intracellular proteins, and cell surface markers.
  • The photobleaching of PE fluorescence was irreversible, allowing for sequential staining and analysis.

Conclusions:

  • This novel method simplifies multi-parameter single-cell analysis by utilizing a single fluorophore and a simple apparatus.
  • The technique expands the potential applications and scope of cytometry.
  • Enables comprehensive analysis of cellular markers and functions without requiring multiple fluorophores.