Robust estimation of bacterial cell count from optical density
Abstract
Optical density (OD) is widely used to estimate the density of cells in liquid culture, but cannot be compared between instruments without a standardized calibration protocol and is challenging to relate to actual cell count. We address this with an interlaboratory study comparing three simple, low-cost, and highly accessible OD calibration protocols across 244 laboratories, applied to eight strains of constitutive GFP-expressing E. coli. Based on our results, we recommend calibrating OD to estimated cell count using serial dilution of silica microspheres, which produces highly precise calibration (95.5% of residuals <1.2-fold), is easily assessed for quality control, also assesses instrument effective linear range, and can be combined with fluorescence calibration to obtain units of Molecules of Equivalent Fluorescein (MEFL) per cell, allowing direct comparison and data fusion with flow cytometry measurements: in our study, fluorescence per cell measurements showed only a 1.07-fold mean difference between plate reader and flow cytometry data.
Additional Information
© The Author(s) 2020. This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/. Received 24 October 2019; Accepted 03 July 2020; Published 17 September 2020. Partial support for this work was provided by NSF Expeditions in Computing Program Award #1522074 as part of the Living Computing Project, and by the Engineering and Physical Sciences Research Council [EP/R034915/1] and EU H2020 [820699]. This document does not contain technology or technical data controlled under either the U.S. International Traffic in Arms Regulations or the U.S. Export Administration Regulations. Data availability: All data generated or analyzed during this study are included in this published article (and its Supplementary Information files). Author Contributions: Conceptualization: J.B., N.G.F., T.H-A., V.S.-1, G.S.B., R.B-T., M.G., D.K., J.M., and C.T.W. Data curation: J.B., N.G.F., T.H-A., and V.S.-1. Formal analysis: J.B. Investigation: Experimental data gathered by iGEM Interlab Study Contributors Methodology: J.B., N.G.F., T.H-A., V.S.-1, G.S.B., R.B-T., M.G., D.K., J.M., V.S.-2, A.S., and C.T.W. Project administration: J.B., N.G.F., and T.H-A. Resources: T.H-A., V.S.-1, and A.S. Software: J.B. Writing (original draft): J.B. and N.G.F. Writing (review & editing): J.B., N.G.F., T.H-A., G.S.B., J.M., C.T.W., and V.S.-2. The authors declare no competing interests.Errata
Beal, J., Farny, N.G., Haddock-Angelli, T. et al. Author Correction: Robust estimation of bacterial cell count from optical density. Commun Biol 3, 640 (2020). https://doi.org/10.1038/s42003-020-01371-9Attached Files
Published - s42003-020-01127-5.pdf
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Supplemental Material - 42003_2020_1127_MOESM1_ESM.pdf
Supplemental Material - 42003_2020_1127_MOESM2_ESM.txt
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Supplemental Material - 42003_2020_1127_MOESM5_ESM.pdf
Supplemental Material - 42003_2020_1127_MOESM6_ESM.pdf
Supplemental Material - 42003_2020_1127_MOESM7_ESM.pdf
Erratum - s42003-020-01371-9.pdf
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Additional details
- PMCID
- PMC7499192
- Eprint ID
- 99256
- Resolver ID
- CaltechAUTHORS:20191014-134916544
- CCF-1522074
- NSF
- EP/R034915/1
- Engineering and Physical Sciences Research Council (EPSRC)
- 820699
- European Research Council (ERC)
- Created
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2019-10-14Created from EPrint's datestamp field
- Updated
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2023-06-01Created from EPrint's last_modified field