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Confocal Microscopy Reveals Cell Surface Receptor Aggregation Through Image Correlation Spectroscopy
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Confocal Microscopy Reveals Cell Surface Receptor Aggregation Through Image Correlation Spectroscopy

Published on: August 2, 2018

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Confocal Microscopy Reveals Cell Surface Receptor Aggregation Through Image Correlation Spectroscopy

Adam C Parslow1, Andrew H A Clayton2, Peter Lock3

  • 1Tumour Targeting Laboratory, Olivia Newton-John Cancer Research Institute; School of Cancer Medicine, La Trobe University.

Journal of Visualized Experiments : Jove
|August 21, 2018
PubMed

Insights

Confocal microscopy quantifies receptor aggregation on cell surfaces using spatial image correlation spectroscopy (ICS). This method aids in developing pre-clinical agents by analyzing fluorescently labeled molecules for drug delivery research.

Area of Science:

  • Cell biology
  • Biophysics
  • Microscopy

Background:

  • Antibody-based cytotoxic drug delivery systems are advancing, necessitating methods to understand their effects on cell surface receptors.
  • Receptor aggregation and internalization are critical processes influenced by these agents, impacting drug efficacy.
  • Characterizing these molecular events is vital for pre-clinical agent development.

Purpose of the Study:

  • To present an accessible protocol for quantifying receptor aggregation on cell surfaces.
  • To enable the characterization of static cell states and potentially expand to temporal investigations.
  • To provide a quantitative measure of target molecule aggregation using established techniques.

Main Methods:

  • Utilizes confocal microscopy and fluorescent immunocytochemistry.
  • Employs the open-source FIJI distribution of ImageJ for image analysis.
  • Performs spatial image correlation spectroscopy (ICS) to quantify fluorescent intensity relative to beam area.

Main Results:

  • Quantifies fluorescent intensity of labeled receptors as a function of confocal microscope beam area.
  • Provides a quantitative measure of target molecule aggregation status on the cell surface.
  • Successfully characterizes receptor clustering events.

Conclusions:

  • Spatial image correlation spectroscopy (ICS) offers an accessible method for quantifying cell surface receptor aggregation.
  • This protocol supports the development of pre-clinical agents by providing insights into molecular interactions.
  • The methodology is adaptable for both static and potentially dynamic cellular investigations.

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