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Related Experiment Videos

Fluorometer based multi-parameter analysis of phagocytic cell activation.

B Seligmann1, K Patel, W O Haston

  • 1Research Department, CIBA-GEIGY Corporation, Summit, NJ 07901.

Agents and Actions
|August 1, 1987
PubMed
Summary
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Neutrophil activation triggers simultaneous calcium rise, membrane potential changes, and shape alterations. Fura 2 measurements reveal lower resting intracellular calcium levels than previously estimated.

Area of Science:

  • Immunology
  • Cellular Biology
  • Biophysics

Background:

  • Neutrophils play a critical role in the immune response.
  • Understanding neutrophil activation pathways is crucial for immunology.
  • Previous studies relied on limited measurements of neutrophil responses.

Purpose of the Study:

  • To simultaneously measure neutrophil responses including calcium flux, membrane potential, and shape change.
  • To investigate the effects of specific neutrophil activators on these parameters.
  • To refine the understanding of intracellular calcium levels in unstimulated neutrophils.

Main Methods:

  • Simultaneous measurement of intracellular calcium concentration using fura 2.
  • Monitoring membrane potential changes.

Related Experiment Videos

  • Assessing cell shape changes via 90-degree light scatter.
  • Activation of neutrophils with diacylglycerol, leukotriene B4, and N-formylmethionyl-leucyl-phenylalanine.

    • Main Results:

    • Simultaneous measurements yield results comparable to independent measurements.
    • Diacylglycerol depolarizes membrane potential and decreases light scatter.
    • Leukotriene B4 induces calcium rise and decreases light scatter.
    • N-formylmethionyl-leucyl-phenylalanine causes membrane depolarization, calcium rise, and decreased light scatter.
    • Fura 2 studies indicate significantly lower intracellular free calcium in unstimulated neutrophils compared to quin 2 estimates.

    Conclusions:

    • Simultaneous monitoring of neutrophil activation markers is feasible and accurate.
    • Specific signaling molecules elicit distinct but overlapping responses in neutrophils.
    • The resting intracellular calcium concentration in neutrophils is lower than previously reported, impacting our understanding of neutrophil signaling.