Statistics of single-molecule measurements: applications in flow-cytometry sizing of DNA fragments

Cytometry. Part a : the Journal of the International Society for Analytical Cytology
Matthew M FerrisR A Keller

Abstract

The measurement of physical properties from single molecules has been demonstrated. However, the majority of single-molecule studies report values based on relatively large data sets (e.g., N > 50). While there are studies that report physical quantities based on small sample sets, there has not been a detailed statistical analysis relating sample size to the reliability of derived parameters. Monte Carlo simulations and multinomial analysis, dependent on quantifiable experimental parameters, were used to determine the minimum number of single-molecule measurements required to produce an accurate estimate of a population mean. Simulation results were applied to the fluorescence-based sizing of DNA fragments by ultrasensitive flow cytometry (FCM). Our simulations show, for an analytical technique with a 10% CV, that the average of as few as five single-molecule measurements would provide a mean value within one SD of the population mean. Additional simulations determined the number of measurements required to obtain the desired number of replicates for each subpopulation within a mixture. Application of these results to flow cytometry data for lambda/HindIII and S. aureus Mu50/SmaI DNA digests produced accurate DNA fingerprints ...Continue Reading

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Citations

Aug 3, 2005·Analytical and Bioanalytical Chemistry·Petra S Dittrich, Andreas Manz
Jan 14, 2009·Journal of Virological Methods·Janet BarlettaNiel T Constantine
Jun 19, 2007·Analytical Biochemistry·Jonas MelinMats Nilsson
Apr 7, 2005·Cytometry. Part a : the Journal of the International Society for Analytical Cytology·Steven W GravesJohn P Nolan
Feb 5, 2005·Analytical Biochemistry·Matthew M FerrisRichard A Keller
Apr 12, 2016·Scientific Reports·Lianhua DongSang-Ryoul Park
Apr 15, 2006·Cytometry. Part a : the Journal of the International Society for Analytical Cytology·James W JacobsonJohn Kettman
May 9, 2006·Cytometry. Part a : the Journal of the International Society for Analytical Cytology·Howard M Shapiro, Nancy G Perlmutter
May 26, 2010·The Analyst·Andrew C BeveridgeThomas M Yoshida

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