Use of Flavin-Related Cellular Autofluorescence to Monitor Processes in Microbial Biotechnology
Abstract
:1. Introduction
2. Materials and Methods
2.1. Microorganisms and Cultivation
2.2. Sample Tubes Preparation for Fluorescence Analysis
2.3. Propidium Iodide Staining Essay for Flow Cytometry
2.4. BODIPY 493/503 Staining Essay
2.5. Flow Cytometry
2.6. Fluorescence Microscopy, FLIM
3. Results
3.1. Analysis of Emission Spectra of Flavins and Whole Bacterial Cells
3.2. Using Green Autofluorescence to Monitor the Viability of Prokaryotic Cell Cultures
3.3. Analysis of Average Fluorescence Lifetimes
4. Discussion
4.1. Analysis of Emission Spectra of Flavins and Whole Bacterial Cells
4.2. Using Green Autofluorescence to Monitor the Viability of Prokaryotic Cell Cultures
4.3. Analysis of Average Fluorescence Lifetimes
4.4. Flavin-PHAs Hypothesis
5. Conclusions
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Conflicts of Interest
References
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Sample | A1 [kCnts] | A2 [kCnts] | τ1 [ns] | τ2 [ns] | A2/A1 | |
---|---|---|---|---|---|---|
Cupriavidus necator H16 | 1 | 0.105 ± 0.004 | 0.197 ± 0.007 | 4.17 ± 0.02 | 1.29 ± 0.06 | 1.9 |
2 | 0.452 ± 0.008 | 0.775 ± 0.023 | 5.04 ± 0.04 | 1.13 ± 0.04 | 1.7 | |
3 | 0.262 ± 0.009 | 0378 ± 0.016 | 4.54 ± 0.05 | 1.22 ± 0.03 | 2.2 | |
4 | 0.279 ± 0.015 | 0.700 ± 0.008 | 4.60 ± 0.12 | 122 ± 0.04 | 2.5 | |
5 | 0.320 ± 0.020 | 0.751 ± 0.012 | 4.30 ± 0.11 | 1.15 ± 0.05 | 2.3 | |
6 | 0.034 ± 0.003 | 0.082 ± 0.004 | 3.80 ± 0.11 | 0.92 ± 0.05 | 2.4 | |
7 | 0.061 ± 0.005 | 0.103 ± 0.003 | 4.l0 ± 0.13 | 1.14 ± 0.07 | 1.7 | |
Cupriavidus necator PHB-4 | 1 | 0359 ± 0.008 | 1.270 ± 0.015 | 4.09 ± 0.04 | 1.12 ± 0.02 | 3.5 |
2 | 0.147 ± 0.006 | 0.479 ± 0.012 | 3.80 ± 0.06 | 1.11 ± 0.03 | 3.3 | |
3 | 0.594 ± 0.012 | 2.420 ± 0.027 | 3.61 ± 0.05 | 1.07 ± 0.01 | 4.1 | |
4 | 0.740 ± 0.020 | 3.210 ± 0.037 | 3.38 ± 0.03 | 1.06 ± 0.02 | 4.3 | |
5 | 0.072 ± 0.003 | 0.220 ± 0.007 | 3.65 ± 0.07 | 0.88 ± 0.03 | 3.1 | |
6 | 0.052 ± 0.006 | 0.161 ± 0.006 | 4.00 ± 0.22 | 0.90 ± 0.06 | 3.1 | |
7 | 0.288 ± 0.015 | 0.979 ± 0.008 | 4.33 ± 0.06 | 1.18 ± 0.03 | 3.4 |
Sample | A1 [kCnts] | A2 [kCnts] | τ1 [ns] | τ2 [ns] | A2/A1 | |
---|---|---|---|---|---|---|
CN H16 | 1 | 6.000 ± 0.018 | 0.549 ± 0.023 | 5.06 ± 0.01 | 1.50 ± 0.15 | 0.09 |
2 | 5.240 ± 0.022 | 0.571 ± 0.017 | 5.11 ± 0.01 | 1.22 ± 0.10 | 0.11 | |
3 | 6.470 ± 0.028 | 0.596 ± 0.022 | 5.19 ± 0.01 | 1.22 ± 0.09 | 0.009 | |
CN PHB-4 | 1 | 0.412 ± 0.019 | 0.476 ± 0.008 | 5.08 ± 0.08 | 1.40 ± 0.11 | 1.15 |
2 | 0.273 ± 0.005 | 0.339 ± 0.007 | 5.07 ± 0.07 | 1.29 ± 0.03 | 1.24 | |
3 | 0.295 ± 0.016 | 0.364 ± 0.016 | 5.10 ± 0.03 | 1.18 ± 0.08 | 1.23 |
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Müllerová, L.; Marková, K.; Obruča, S.; Mravec, F. Use of Flavin-Related Cellular Autofluorescence to Monitor Processes in Microbial Biotechnology. Microorganisms 2022, 10, 1179. https://doi.org/10.3390/microorganisms10061179
Müllerová L, Marková K, Obruča S, Mravec F. Use of Flavin-Related Cellular Autofluorescence to Monitor Processes in Microbial Biotechnology. Microorganisms. 2022; 10(6):1179. https://doi.org/10.3390/microorganisms10061179
Chicago/Turabian StyleMüllerová, Lucie, Kateřina Marková, Stanislav Obruča, and Filip Mravec. 2022. "Use of Flavin-Related Cellular Autofluorescence to Monitor Processes in Microbial Biotechnology" Microorganisms 10, no. 6: 1179. https://doi.org/10.3390/microorganisms10061179