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Imaging Biological Samples with Optical Microscopy

Imaging Biological Samples with Optical Microscopy

Optical microscopy uses optic principles to provide detailed images of samples. Antonie van Leeuwenhoek designed the first compound optical microscope in the 17th century to visualize blood cells, bacteria, and yeast cells. In 1830, Joseph Jackson Lister created an essentially modern light microscope. The 20th century saw the development of microscopes with enhanced magnification and resolution.
In optical microscopy, the specimen to be viewed is placed on a glass slide and clipped on the stage...

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Updated: Jun 17, 2026

Mapping Absolute DNA Density in Cell Nuclei using Single-molecule Localization Microscopy

Mapping Absolute DNA Density in Cell Nuclei using Single-molecule Localization Microscopy

Published on: November 11, 2025

HOW ACCURATELY CAN A SINGLE MOLECULE BE LOCALIZED WHEN IMAGED THROUGH AN OPTICAL MICROSCOPE?

Sripad Ram¹, E Sally Ward, Raimund J Ober

¹Center for Immunology, University of Texas Southwestern Medical Center, Dallas, TX.

Proceedings. IEEE International Symposium on Biomedical Imaging

|September 28, 2011

Summary

We derived a formula to determine the ultimate accuracy for pinpointing single molecules using optical microscopy. This limit is influenced by microscope optics and molecule properties, and is worsened by detector pixelation and noise.

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Last Updated: Jun 17, 2026

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Published on: December 9, 2013

Area of Science:

Optical microscopy
Single-molecule imaging
Nanoscale localization

Background:

Accurate localization of single molecules is crucial for various scientific fields.
Current methods face limitations in precision due to optical and detection constraints.

Purpose of the Study:

To derive a fundamental expression for the limit of single-molecule localization accuracy.
To identify key factors influencing this limit in optical microscopy.

Main Methods:

Development of a simple analytical expression.
Analysis of optical and photophysical properties.
Inclusion of detector pixelation and noise effects.

Main Results:

An analytical expression for the fundamental localization limit was established.
The impact of microscope optics and molecular photophysics on accuracy was quantified.
The detrimental effects of pixelation and noise were demonstrated.

Conclusions:

The derived expression provides a benchmark for achievable single-molecule localization accuracy.
Experimental design can be optimized by understanding the factors limiting localization precision.
This work offers practical insights for improving single-molecule imaging experiments.