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Techniques & Tools Spectroscopy, Education

Landmark Literature: Spectroscopy

A Single Step

Norman Dovichi

Landmark Paper: DY Chen and NJ Dovichi, “Singlemolecule detection in capillary electrophoresis: Molecular shot noise as a fundamental limit to chemical analysis”, Anal Chem, 68, 690-696 (1996).

Analytical chemists spent most of the 20th century developing and improving instrumentation to perform ultrasensitive chemical analysis. The community ultimately achieved single molecule detection in the 1980s with laser-induced fluorescence. Those early studies were simply interested in addressing the formidable issues associated with minimizing noise, maximizing signal, and processing data to confidently detect and count molecules. This paper with David Chen was the first to discuss fundamental ramifications of measurements with small numbers of molecules.

Optical shot noise – observed as a Poisson distribution in detected photons in photon counting experiments – is a fundamental noise source that ultimately limits the precision of spectroscopic measurements. In this paper, we demonstrated the analogous phenomenon of molecular shot-noise, where performing analysis on small numbers of analyte molecules introduces a fundamental noise source. As expected, molecular shot noise limits the precision of quantitative measurements, particularly when dealing with small numbers of molecules. It was more surprising to discover that molecular shot noise also limited the precision with which we can measure migration time and peak width in separations.

What makes it stand out? This paper was the first systematic study of fundamental issues associated with quantitative analysis on small numbers of molecules. As analytical chemists, we have been trained to consider the accuracy and precision of measurements. Single molecule counting is the ultimate quantitative measurement; after all, one can have no higher accuracy than counting the number of molecules in a sample. However, the precision of that measurement is ultimately limited by Poisson statistics: fluctuations in the number of molecules taken for chemical analysis is a fundamental and irreducible source of uncertainty.

Our experience with ultrasensitive analysis based on fluorescence detection with capillary electrophoresis informed our work on separation of DNA fragments generated by Sanger sequencing, which resulted in our development of capillary array electrophoresis for highthroughput DNA analysis, which led to the sequencing of the human genome ahead of schedule and under budget.

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About the Authors

Norman Dovichi

Norm is the Grace-Rupley Professor of Chemistry and Biochemistry, at the University of Notre Dame, Indiana, USA. His postdoctoral fellowship at Los Alamos Scientific Lab introduced the concept of single molecule detection, leading to the development of a capillary array DNA sequencer that became the workhorse tool used in the human genome project.


Mary Kate Donais

Mary Kate Donais is Professor in the Department of Chemistry at St Anselm College, New Hampshire, USA, and has worked as an analytical chemist in federal government, industrial, and academic settings. She is Chair of 2016’s SciX conference. 


Marcia F Mesko

Associate Professor, Center for Chemical, Pharmaceutical and Food Sciences, Federal University of Pelotas, Pelotas, Brazil.


Dominic J. Hare

Associate Professor and Principal Research Fellow, Atomic Pathology Laboratory, Melbourne and Dementia Research Centre at the Florey Institute of Neuroscience and Mental Health and the University of Melbourne, Australia.

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