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Confocal Fluorescence Microscopy

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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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Related Experiment Video

Updated: May 14, 2026

Single Molecule Fluorescence Microscopy on Planar Supported Bilayers

Single Molecule Fluorescence Microscopy on Planar Supported Bilayers

Published on: October 31, 2015

Multi-beam confocal microscopy based on a custom image sensor with focal-plane pinhole array effect.

Keiichiro Kagawa¹, Min-Woong Seo, Keita Yasutomi

¹Research Institute of Electronics, Shizuoka University, 3-5-1 Johoku, Naka-ku, Hamamatsu, Shizuoka 432-8011, Japan. kagawa@idl.rie.shizuoka.ac.jp

|February 8, 2013

Summary

A novel multi-beam confocal microscopy system eliminates the need for a physical pinhole. This was achieved using a custom CMOS image sensor, demonstrating successful confocality with an axial resolution of 8.9 μm.

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Area of Science:

Optics and Photonics
Microscopy Technology
Image Sensors

Background:

Confocal microscopy offers high resolution but typically requires a physical pinhole.
Traditional pinholes can limit light throughput and system complexity.

Purpose of the Study:

To demonstrate multi-beam confocal microscopy without a physical pinhole.
To develop and validate a custom CMOS image sensor for this application.

Main Methods:

Developed a custom CMOS image sensor with special pixel addressing.
Implemented photocarrier discarding to simulate a focal-plane pinhole array effect.
Configured a multi-beam illumination system.

Main Results:

Achieved confocal microscopy operation without a physical pinhole.
Measured an axial resolution of 8.9 μm (FWHM) using a planar mirror.
Validated the confocality of the proposed CMOS image sensor.

Conclusions:

The custom CMOS image sensor effectively replaces the physical pinhole in confocal microscopy.
This approach enables simplified and potentially more efficient multi-beam confocal imaging.
The demonstrated technology paves the way for advanced microscopy techniques.