Cellular and Molecular Life Sciences

, Volume 72, Issue 2, pp 367–381 | Cite as

Imaging oxygen in neural cell and tissue models by means of anionic cell-permeable phosphorescent nanoparticles

  • Ruslan I. DmitrievEmail author
  • Sergey M. Borisov
  • Alina V. Kondrashina
  • Janelle M. P. Pakan
  • Ujval Anilkumar
  • Jochen H. M. Prehn
  • Alexander V. Zhdanov
  • Kieran W. McDermott
  • Ingo Klimant
  • Dmitri B. Papkovsky
Research Article


Cell-permeable phosphorescent probes enable the study of cell and tissue oxygenation, bioenergetics, metabolism, and pathological states such as stroke and hypoxia. A number of such probes have been described in recent years, the majority consisting of cationic small molecule and nanoparticle structures. While these probes continue to advance, adequate staining for the study of certain cell types using live imaging techniques remains elusive; this is particularly true for neural cells. Here we introduce novel probes for the analysis of neural cells and tissues: negatively charged poly(methyl methacrylate-co-methacrylic acid)-based nanoparticles impregnated with a phosphorescent Pt(II)-tetrakis(pentafluorophenyl)porphyrin (PtPFPP) dye (this form is referred to as PA1), and with an additional reference/antennae dye poly(9,9-diheptylfluorene-alt-9,9-di-p-tolyl-9H-fluorene) (this form is referred to as PA2). PA1 and PA2 are internalised by endocytosis, result in efficient staining in primary neurons, astrocytes, and PC12 cells and multi-cellular aggregates, and allow for the monitoring of local O2 levels on a time-resolved fluorescence plate reader and PLIM microscope. PA2 also efficiently stains rat brain slices and permits detailed O2 imaging experiments using both one and two-photon intensity-based modes and PLIM modes. Multiplexed analysis of embryonic rat brain slices reveals age-dependent staining patterns for PA2 and a highly heterogeneous distribution of O2 in tissues, which we relate to the localisation of specific progenitor cell populations. Overall, these anionic probes are useful for sensing O2 levels in various cells and tissues, particularly in neural cells, and facilitate high-resolution imaging of O2 in 3D tissue models.


Biomaterials Cell and tissue oxygen Intracellular oxygen probe Multiplexed detection Nanosensors Phosphorescence quenching microscopy PLIM 





Brain lipid-binding protein




Cholera toxin, subunit B


Days in vitro




Dimethyl sulfoxide




Green fluorescent protein


Hanks balanced salt solution


Hoechst 33342


Intracellular O2






Polyacrylate NP


Phosphate buffered saline




Photodynamic therapy


Phosphorescence lifetime imaging microscopy


Poly(methyl methacrylate-co-methacrylic acid






Region of interest


Room temperature


Tris-buffered saline, tween 20


Time-correlated single photon counting


Time-resolved fluorescence



This work was supported by the Science Foundation Ireland, Grant 12/RC/2276, the European Commission FP7 Program, grant FP7-HEALTH-2012-INNOVATION-304842-2, the Irish Research Council for Science, Engineering and Technology, the Health Research Board, the Programme for Research at Third Level Institutions Cycle 4 and Marie Curie IAPP Oxy-Sense (No. 230641). We thank T. Foley and Dr. Y. Nolan (Department of Anatomy and Neuroscience, UCC) for the help with primary neuronal cultures and Dr. Heiko Dussmann (Royal College of Surgeons in Ireland, Dublin) for help with microscopy imaging.

Supplementary material

18_2014_1673_MOESM1_ESM.pdf (493 kb)
Supplementary material 1 (PDF 492 kb)


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Copyright information

© Springer Basel 2014

Authors and Affiliations

  • Ruslan I. Dmitriev
    • 1
    Email author
  • Sergey M. Borisov
    • 2
  • Alina V. Kondrashina
    • 1
  • Janelle M. P. Pakan
    • 3
  • Ujval Anilkumar
    • 4
  • Jochen H. M. Prehn
    • 4
  • Alexander V. Zhdanov
    • 1
  • Kieran W. McDermott
    • 3
  • Ingo Klimant
    • 2
  • Dmitri B. Papkovsky
    • 1
  1. 1.School of Biochemistry and Cell BiologyUniversity College CorkCorkIreland
  2. 2.Institute of Analytical Chemistry and Food ChemistryGraz University of TechnologyGrazAustria
  3. 3.Department of Anatomy and NeuroscienceUniversity College CorkCorkIreland
  4. 4.Department of Physiology and Medical Physics, Centre for the Study of Neurological DisordersRoyal College of Surgeons in IrelandDublinIreland

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