Oxygen sensor nanoparticles for monitoring bacterial growth and characterization of dose–response functions in microfluidic screenings

Abstract

We are presenting a microfluidic droplet-based system for non-invasive, simultaneous optical monitoring of oxygen during bacterial cultivation in nL-sized droplets using ~350 nm nanobeads made from polystyrene and doped with the NIR-emitting oxygen probe platinum (II) 5, 10, 15, 20-meso-tetraphenyltetrabenzoporphyrin (PtTPTBP). Data were readout by a two-channel micro flow-through fluorimeter and a two-channel micro flow-through photometer. The time-resolved miniaturized optical multi endpoint detection was applied to simultaneously sense dissolved oxygen, cellular autofluorescence, and cell density in nL-sized segments. Two bacterial strains were studied that are resistant to heavy metal ions, viz. Streptomyces acidiscabies E13 and Psychrobacillus psychrodurans UrPLO1. The study has two main features in that it demonstrates (a) the possibility to monitor the changes in oxygen partial pressure during metabolic activity of different bacterial cultures inside droplets, and (b) the efficiency of droplet-based microfluidic techniques along with multi-parameter optical sensing for highly resolved microtoxicological screenings in aquatic systems.

Microfluidic droplet-based system with multi-parameter optical sensing for bacterial cultivation and highly resolved microtoxicological screenings in nanoliter droplets.

This is a preview of subscription content, access via your institution.

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5

References

  1. 1.

    Dusenbery DB (1996) Life at small scale—the behavior of microbes. Scientific American Library, New York

    Google Scholar 

  2. 2.

    Woese CR (1987) Bacterial evolution. Microbiol Rev 51:221–271

    CAS  Google Scholar 

  3. 3.

    Pace NR (1997) A molecular view of microbial diversity and the biosphere. Science 276:734–740

    CAS  Article  Google Scholar 

  4. 4.

    Ward DM, Weller R, Bateson MM (1990) 16s ribosomal-Rna sequences reveal numerous uncultured microorganisms in a natural community. Nature 345:63–65

    CAS  Article  Google Scholar 

  5. 5.

    Haferburg G, Kothe E (2007) Microbes and metals: interactions in the environment. J Basic Microbiol 47:453–467

    CAS  Article  Google Scholar 

  6. 6.

    Vallenet D, Engelen S, Mornico D, Cruveiller S, Fleury L, Lajus A, Rouy Z, Roche D, Salvignol G, Scarpelli C, Medigue C (2009) MicroScope: a platform for microbial genome annotation and comparative genomics. Database-Oxford

  7. 7.

    Suthanthiran M (1997) Clinical application of molecular biology: a study of allograft rejection with polymerase chain reaction. Am J Med Sci 313:264–267

    CAS  Article  Google Scholar 

  8. 8.

    Joo C, Balci H, Ishitsuka Y, Buranachai C, Ha T (2008) Advances in single-molecule fluorescence methods for molecular biology. Annu Rev Biochem 77:51–76

    CAS  Article  Google Scholar 

  9. 9.

    Stone HA, Stroock AD, Ajdari A (2004) Engineering flows in small devices: microfluidics toward a lab-on-a-chip. Annu Rev Fluid Mech 36:381–411

    Article  Google Scholar 

  10. 10.

    Budden M, Schneider S, Groß A, Kohler JM (2013) Microfluidic encoding: generation of arbitrary droplet sequences by electrical switching in microchannels. Sensors Actuators A Phys 189:288–297

    CAS  Article  Google Scholar 

  11. 11.

    Yang CG, Xu ZR, Wang JH (2010) Manipulation of droplets in microfluidic systems. TrAC Trends Anal Chem 29:141–157

    CAS  Article  Google Scholar 

  12. 12.

    Cao J, Kursten D, Schneider S, Knauer A, Gunther PM, Kohler JM (2012) Uncovering toxicological complexity by multi-dimensional screenings in microsegmented flow: modulation of antibiotic interference by nanoparticles. Lab Chip 12:474–484

    CAS  Article  Google Scholar 

  13. 13.

    Lim CT, Zhang Y (2007) Bead-based microfluidic immunoassays: the next generation. Biosens Bioelectron 22:1197–1204

    CAS  Article  Google Scholar 

  14. 14.

    Hartung R, Brosing A, Sczcepankiewicz G, Liebert U, Hafner N, Durst M, Felbel J, Lassner D, Kohler JM (2009) Application of an asymmetric helical tube reactor for fast identification of gene transcripts of pathogenic viruses by micro flow-through PCR. Biomed Microdevices 11:685–692

    CAS  Article  Google Scholar 

  15. 15.

    Funfak A, Cao JL, Wolfbeis O, Martin K, Kohler J (2009) Monitoring cell cultivation in microfluidic segments by optical pH sensing with a micro flow-through fluorometer using dye-doped polymer particles. Microchim Acta 164:279–286

    CAS  Article  Google Scholar 

  16. 16.

    Semenza GL (2007) Life with oxygen. Science 318:62–64

    CAS  Article  Google Scholar 

  17. 17.

    Winkler LW (1888) Die Bestimmung des im Wasser gelösten Sauerstoffes. Ber Dtsch Chem Ges 21:2843–2854

    Article  Google Scholar 

  18. 18.

    Kinoshita K (1992) Electrochemical oxygen technology. In: Wiley-Interscience

  19. 19.

    Vanderkooi JM, Maniara G, Green TJ, Wilson DF (1987) An optical method for measurement of dioxygen concentration based upon quenching of phosphorescence. J Biol Chem 262:5476–5482

    CAS  Google Scholar 

  20. 20.

    Stich MIJ, Schaeferling M, Wolfbeis OS (2009) Multicolor fluorescent and permeation-selective microbeads enable simultaneous sensing of pH, oxygen, and temperature. Adv Mater 21:2216–2220

    CAS  Article  Google Scholar 

  21. 21.

    Nagl S, Stich MIJ, Schaferling M, Wolfbeis OS (2009) Method for simultaneous luminescence sensing of two species using optical probes of different decay time, and its application to an enzymatic reaction at varying temperature. Anal Bioanal Chem 393:1199–1207

    CAS  Article  Google Scholar 

  22. 22.

    Borisov SM, Klimant I (2009) Luminescent nanobeads for optical sensing and imaging of dissolved oxygen. Microchim Acta 164:7–15

    CAS  Article  Google Scholar 

  23. 23.

    Abbyad P, Tharaux PL, Martin JL, Baroud CN, Alexandrou A (2010) Sickling of red blood cells through rapid oxygen exchange in microfluidic drops. Lab Chip 10:2505–2512

    CAS  Article  Google Scholar 

  24. 24.

    Wang XD, Achatz DE, Hupf C, Sperber M, Wegener J, Bange S, Lupton JM, Wolfbeis OS (2013) Imaging of cellular oxygen via two-photon excitation of fluorescent sensor nanoparticles. Sensors Actuators B Chem 188:257–262

    Article  Google Scholar 

  25. 25.

    Stitt DT, Nagar MS, Haq TA, Timmins MR (2002) Determination of growth rate of microorganisms in broth from oxygen-sensitive fluorescence plate reader measurements. Biotechniques 32:684

    CAS  Google Scholar 

  26. 26.

    Jasionek G, Ogurtsov V, Papkovsky D (2013) Rapid detection and respirometric profiling of aerobic bacteria on panels of selective media. J Appl Microbiol 114:423–432

    CAS  Article  Google Scholar 

  27. 27.

    O'Mahony FC, Papkovsky DB (2006) Rapid high-throughput assessment of aerobic bacteria in complex samples by fluorescence-based oxygen respirometry. Appl Environ Microbiol 72:1279–1287

    Article  Google Scholar 

  28. 28.

    Arain S, John GT, Krause C, Gerlach J, Wolfbeis OS, Klimant I (2006) Characterization of microtiterplates with integrated optical sensors for oxygen and pH, and their applications to enzyme activity screening, respirometry, and toxicological assays. Sensors Actuators B Chem 113:639–648

    CAS  Article  Google Scholar 

  29. 29.

    O'Mahony FC, O'Donovan C, Hynes J, Moore T, Davenport J, Papkovsky DB (2005) Optical oxygen microrespirometry as a platform for environmental toxicology and animal model studies. Environ Sci Technol 39:5010–5014

    Article  Google Scholar 

  30. 30.

    Hopwood DA (2006) Soil to genomics: the streptomyces chromosome. Annu Rev Genet 40:1–23

    CAS  Article  Google Scholar 

  31. 31.

    Valko M, Morris H, Cronin MTD (2005) Metals, toxicity and oxidative stress. Curr Med Chem 12:1161–1208

    CAS  Article  Google Scholar 

  32. 32.

    Langella F, Grawunder A, Stark R, Weist A, Merten D, Haferburg G, Buchel G, Kothe E (2013) Microbially assisted phytoremediation approaches for two multi-element contaminated sites. Environ Sci Pollut Res Int. doi:10.1007/s11356-11013-12165-11350

    Google Scholar 

  33. 33.

    Rogers JE, Nguyen KA, Hufnagle DC, McLean DG, Su WJ, Gossett KM, Burke AR, Vinogradov SA, Pachter R, Fleitz PA (2003) Observation and interpretation of annulated porphyrins: studies on the photophysical properties of meso-tetraphenylmetalloporphyrins. J Phys Chem A 107:11331–11339

    CAS  Article  Google Scholar 

  34. 34.

    Borisov SM, Nuss G, Klimant I (2008) Red light-excitable oxygen sensing materials based on platinum(II) and palladium(II) benzoporphyrins. Anal Chem 80:9435–9442

    CAS  Article  Google Scholar 

  35. 35.

    Cao J, Kursten D, Krause K, Kothe E, Martin K, Roth M, Kohler JM (2013) Application of micro-segmented flow for two-dimensional characterization of the combinatorial effect of zinc and copper ions on metal-tolerant Streptomyces strains. Appl Microbiol Biotechnol 97:8923–8930

    CAS  Article  Google Scholar 

  36. 36.

    Preininger C, Klimant I, Wolfbeis OS (1994) Optical-fiber sensor for biological oxygen-demand. Anal Chem 66:1841–1846

    CAS  Article  Google Scholar 

  37. 37.

    Riedel K, Renneberg R, Kuhn M, Scheller F (1988) A fast estimation of biochemical oxygen-demand using microbial sensors. Appl Microbiol Biotechnol 28:316–318

    CAS  Google Scholar 

  38. 38.

    Papkovsky DB, Dmitriev RI (2013) Biological detection by optical oxygen sensing. Chem Soc Rev 42:8700–8732

    CAS  Article  Google Scholar 

  39. 39.

    John GT, Klimant I, Wittmann C, Heinzle E (2003) Integrated optical sensing of dissolved oxygen in microtiter plates: a novel tool for microbial cultivation. Biotechnol Bioeng 81:829–836

    CAS  Article  Google Scholar 

  40. 40.

    Lam RHW, Kim MC, Thorsen T (2009) Culturing aerobic and anaerobic bacteria and mammalian cells with a microfluidic differential oxygenator. Anal Chem 81:5918–5924

    CAS  Article  Google Scholar 

Download references

Acknowledgments

J. Cao is funded by German Federal Environmental Foundation under contract No. 20009/009. S. Nagl is funded by and thanks the German Research Foundation (DFG, NA 947/1-2). The authors gratefully acknowledge the funding from German Federal Ministry of Education and Research in the frame of the project BactoCat (Kz: 031A161A). We thank S. Schneider, F. Weise and F. Möller for excellent experimental and technical assistance, Prof. N. Sträter and Dr. C. Roth (Institute of Bioanalytical Chemistry, University of Leipzig) for dynamic light scattering and Dr. A. Funfak (Laboratoire d'Hydrodynamique, Ecole Polytechnique, Palaiseau, France) for helpful discussions.

Author information

Affiliations

Authors

Corresponding author

Correspondence to Jialan Cao.

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Cao, J., Nagl, S., Kothe, E. et al. Oxygen sensor nanoparticles for monitoring bacterial growth and characterization of dose–response functions in microfluidic screenings. Microchim Acta 182, 385–394 (2015). https://doi.org/10.1007/s00604-014-1341-3

Download citation

Keywords

  • Droplet-based microfluidics
  • Oxygen sensor
  • Optical probe-doped micro and nanoparticles
  • Micro cultivation
  • Highly-resolved dose/response function