Microchimica Acta

, 186:529 | Cite as

Absolute quantification of particle number concentration using a digital single particle counting system

  • Li Zhang
  • Yu-Jun Yang
  • Jun-Yi Xiong
  • Zhen Wu
  • Zhi-Xiong Xie
  • Dai-Wen Pang
  • Zhi-Ling ZhangEmail author
Original Paper


The accurate determination of the molar concentration or the number concentration of particles in a defined volume is important but challenging. Since particle diversity and heterogeneity cannot be ignored in particle quantification, single particle counting has become quite important. However, most methods require standard samples (calibrators) which are usually difficult to obtain. The authors describe a method for single particle counting that is based on the combination of digital counting and formation of microdroplets in a microchip. By compartmentalizing particles into picoliter droplets, positive droplets encapsulating particles were counted and particle concentrations were calculated by Poisson statistics. The concentration of particles over a wide range (from 5.0 × 103 to 1.8 × 107 particles per mL) were accurately determined without the need for using a calibrator. A microdroplet chip including a T-junction channel achieved a 9-fold increase of signal-to-background ratio compared to the traditional flow-focusing chip. This makes the digital counting system a widely applicable tool for quantification of fluorescent particles. Various particles including differently sized fluorescent microspheres and bacteria with large heterogeneity in shape such as Escherichia coli DH5α-pDsRed were accurately quantified by this method.

Graphical abstract

Schematic representation of the digital single particle counting system for absolute quantification of particles. Particles compartmentalized in picoliter droplets were counted and the number concentration of particles was determined using digital analysis.


Digital analysis Number concentration determination Poisson statistics Droplet microfluidics Picoliter droplet 



This work was supported by the National Natural Science Foundation of China (21775111), and the National Science and Technology Major Project of China (2018ZX10301405).

Compliance with ethical standards

The author(s) declare that they have no competing interests.

Supplementary material

604_2019_3692_MOESM1_ESM.docx (19.6 mb)
ESM 1 (DOCX 20108 kb)


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Copyright information

© Springer-Verlag GmbH Austria, part of Springer Nature 2019

Authors and Affiliations

  1. 1.Key Laboratory of Analytical Chemistry for Biology and Medicine (Ministry of Education), College of Chemistry and Molecular Sciences, State Key Laboratory of VirologyWuhan UniversityWuhanPeople’s Republic of China
  2. 2.College of Life SciencesWuhan UniversityWuhanPeople’s Republic of China

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