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Confocal Fluorescence Microscopy

Confocal Fluorescence Microscopy

Confocal microscopy is an advanced microscopic technique. The prime advantage of the confocal microscope over other microscopy techniques is its ability to block the out-of-focus light from the illuminated samples using pinholes. It is widely used with fluorescence optics to obtain high-resolution, sharp contrast images. Unlike optical microscopes, confocal microscopes use a focused beam of light laser to scan the entire sample surface at different z-planes. These microscopes are, therefore,...

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Confocal Microscopy Reveals Cell Surface Receptor Aggregation Through Image Correlation Spectroscopy

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Practical considerations in acquiring biological signals from confocal microscope: solvent effect and temperature

M F Lee¹, S K Kong, K P Fung

¹Department of Biochemistry, Chinese University of Hong Kong, Shatin.

Biological Signals

|September 1, 1996

Summary

Fluorescence microscopic imaging (FMI) can produce artifacts. This study identifies DMSO water content and temperature-induced refractive index changes as sources of false signals in cellular studies.

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Last Updated: Jun 30, 2026

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Area of Science:

Cell Biology
Microscopy Techniques

Background:

Fluorescence microscopic imaging (FMI) is crucial for studying live single-cell dynamics.
Intracellular messengers like Ca2+, H+, and cAMP are commonly monitored using FMI.

Purpose of the Study:

To identify and describe artifacts in confocal scanning fluorescence microscopy.
To prevent misinterpretation of experimental data in cellular studies.

Main Methods:

Confocal scanning fluorescence microscopy was employed.
Cellular responses were monitored to detect anomalies.
Experimental conditions were analyzed to pinpoint artifact origins.

Main Results:

DMSO water content can impact indicator loading and create spurious fluorescence spots.
Temperature fluctuations altering immersion oil refractive index cause focal plane shifts, mimicking spontaneous cellular oscillations.

Conclusions:

Two significant artifact sources in FMI were identified: DMSO quality and temperature-dependent optical effects.
Recommendations are provided for controlling and eliminating these false images in cellular research.