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Related Experiment Videos

Signal amplification in flow cytometry using biotin tyramine.

J C Earnshaw1, J K Osbourn

  • 1Cambridge Antibody Technology, Melbourn, Cambridgeshire, United Kingdom.

Cytometry
|December 20, 1999
PubMed
Summary

Catalysed reporter deposition (CARD) enhances flow cytometry signals by amplifying horseradish peroxidase (HRP) activity. This method significantly boosts detection sensitivity for cell surface markers like CD4 and CD36.

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Area of Science:

  • Immunology
  • Biotechnology
  • Analytical Chemistry

Background:

  • Catalysed reporter deposition (CARD) is a signal amplification technique used in immunoassays.
  • It utilizes horseradish peroxidase (HRP)-conjugated reagents and biotin tyramine for signal enhancement.
  • Previous applications were limited to solid-phase assays.

Purpose of the Study:

  • To demonstrate the application of CARD for signal enhancement in flow cytometry.
  • To evaluate its effectiveness in amplifying signals for cell surface marker detection.

Main Methods:

  • Applied CARD technique to flow cytometry analysis.
  • Used anti-human CD4 and anti-human CD36 antibodies for detection on human lymphocytes and mononuclear cells.
  • Optimized biotin tyramine concentrations and evaluated signal amplification.

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Main Results:

  • Achieved a fivefold signal increase for anti-human CD4 detection compared to standard methods.
  • Repeated biotin tyramine treatment for anti-CD36 resulted in an additional 2.5-fold signal amplification.
  • Demonstrated successful signal enhancement in flow cytometry.

Conclusions:

  • The CARD technique effectively amplifies signals in flow cytometry.
  • This method offers a valuable tool for improving the sensitivity of flow cytometry detection reagents.