Abstract
This article is devoted to the development of a measurement procedure of the cell number concentration in liquids. This procedure is intended to determine the number of viable microorganisms, such as Bifidobacterium, Lactobacillus acidophilus, Listeria monocytogenes, Cronobacter sakazakii, Clostridium perfringens, Staphylococcus aureus, Proteus, Enterococcus, coliforms, Vibrio parahaemolyticus, and Bacillus cereus, in liquids. This methodology implements two main measurement methods: the Petri dish method and direct counting using an inverted fluorescence microscope. It describes the conditions and procedures that ensure measurement accuracy, the scope of the methodology, and the new approaches that enabled the development of a methodology with the best measurement accuracy available for these methods.
The established measurement range for cell number concentration varies from 1 to 1 · 105 1/cm3, allowing the methodology to be applied to a variety of samples, including both highly diluted and concentrated solutions.
The methodology’s application areas include healthcare, veterinary medicine, and the food industry, making it an important tool for quality control of laboratory analyses and food safety assessments. The developed methodology can serve as the basis for the creation of reference materials for cell number concentration, which will facilitate the standardization of measurement processes and increase confidence in the analysis results among consumers and regulatory authorities.
This article makes a significant contribution to the development of measurement methods in microbiology and can be useful for both scientific research and practical applications in various fields.
Keywords
liquid cell counting method, volumetric seeding method, direct counting method using an inverted fluorescence microscope
References
-
Doshi P. The elusive definition of pandemic influenza // Bulletin of the World Health Organization. 2011. V. 89. № 7. P. 532–538.
https://doi.org/10.2471/BLT.11.086173.
-
Kraemer M. UG, Sinka M.E, Duda K.A. et al. The global distribution of the arbovirus vectors Aedes aegypti and Ae. albopictus // Elife. 2015 Jun 30. 4. e08347.
https://doi.org/10.7554/eLife.08347.
-
Microbial Risk Management in Food Processes. 2nd Workshop Arranged by SAFOODNET — Food Safety and Hygiene Networking within New Member States and Assosiated Candidate Countries. VTT Symposium 260. Lyngby, Denmark, October 13–15, 2008 / G. Wirtanen, S. Salo (eds.). Espoo: VTT Technical Research Centre of Finland, 2009. 100 p.
-
Weidi Xie, Noble J.A., Zisserman A. Microscopy cell counting and detection with fully convolutional regression networks // Computer Methods in Biomechanics and Biomedical Engineering: Imaging & Visualization, 2016. P. 283–292.
https://doi.org/10.1080/21681163.2016.1149104.
-
Jeremias J., Pape C. Stamped counting for biomedical images // Methods in Microscopy. 2025. V. 2. № 2. P. 203–213.
https://doi.org/10.1515/mim-2024-0021.
-
Mohammed A.A., Fonder C., Ying Wei, Tavanapong W., Sakaguchi D.S., Qi Li, Mallapragada S.K. CellFMCount: a fluorescence microscopy dataset, benchmark, and methods for cell counting // Computer Vision and Pattern Recognition (cs.CV). The IEEE International Conference on Data Mining (ICDM). November 12–15, 2025, Washington, D.C., USA. IEEE, 2025.
https://doi.org/10.1109/ICDM65498.2025.00069.
-
Mingyue Xiao, Ronghua Luo, Qinghua Liang et al. Anemoside B4 inhibits SARSCoV- 2 replication in vitro and in vivo // Chinese Herbal Medicines. 2024. V. 16. № 1. P. 106–112.
https://doi.org/10.1016/j.chmed.2023.09.005.
-
Paltiel A.D., Zheng A., Walensky R.P. Assessment of SARS-CoV-2 screening strategies to permit the safe reopening of college campuses in the United States // JAMA Network Open. 2020. V. 3. № 7. e2016818.
https://doi.org/10.1001/jamanetworkopen.2020.16818.
-
Miller M.J., Dasgupta Sh., Ruffin J. et al. Severe acute respiratory syndrome coronavirus 2 testing trends among persons aged // Journal of Adolescent Health. 2021. V. 69. № 1. P. 144–148.
https://doi.org/10.1016/j.jadohealth.2021.04.010.
-
Graham B.S., Sullivan N.J. Emerging viral diseases from a vaccinology perspective: preparing for the next pandemic // Nature Immunology. 2018. V. 19. № 1. P. 20–28.
https://doi.org/10.1038/s41590-017-0007-9.
-
Excler J.-L., Saville M., Berkley S., Kim J.H. Vaccine development for emerging infectious diseases // Nature Medicine. 2021. V. 27. № 4. P. 591–600.
https://doi.org/10.1038/s41591-021-01301-0.
-
Aladhadh M. A review of modern methods for the detection of foodborne pathogens // Microorganisms. 2023. V. 11. № 5. 1111.
https://doi.org/10.3390/microorganisms11051111.
For citation
Kulyabina E.V., Melkova O.N., Kulyabina T.V., Dorman M.V., Bulygin F.V., Kirilyuk A.A., Morozova V.V., Morozov V.Yu. Development of a method for measuring cell number concentration in liquid. Almanac of Modern Metrology. 2026; 46 (2): 135–148.
Financing
The authors declare that they received no external financial support for this study.
Contribution of the authors
E.V. Kulyabina developed the concept, formulated the key goals and objectives, implemented the work, obtained the measurement results, and drafted the manuscript and the first version of the article. F.V. Bulygin provided scientific supervision of the project. T.V. Kulyabina conducted the research, including experiments, analysis, and interpretation of all obtained data. O.N. Melkova conducted the research using high-precision laboratory equipment. V.V. Morozova developed the study design. M.V. Dorman implemented the microscope. V.Yu. Morozov applied statistical and mathematical methods for data analysis and performed the data analysis. A.A. Kirilyuk obtained the measurement results. All authors contributed to the writing of the article and participated in its revision. All authors read and approved the final manuscript.
Conflict of interest
The authors declare that they have no potential conflict of interest related to the research presented in this article.
This work is licensed under a Creative Commons Attribution 4.0 License (CC BY 4.0)
©
Elena V. Kulyabina, Olga N. Melkova, Tatiana V. Kulyabina, Maria V. Dorman, Fedor V. Bulygin, Alexander A. Kirilyuk, Valeriia V. Morozova, Vitaliy Yu. Morozov, 2026
JATS XML
This work is available at Scientific Electronic Library eLIBRARY.
Subscribe print journal.
