Evaluation of the derivates of phosphorescent Pt-coproporphyrin as intracellular oxygen-sensitive probes

• Published in: Analytical and bioanalytical chemistry 2010-03, Vol.396 (5), p.1793-1803
• Main Authors: Fercher, Andreas, Ponomarev, Gelii V, Yashunski, Dmitri, Papkovsky, Dmitri
• Format: Article
• Language: English
• Subjects: Analytical Chemistry; Animals; Biochemistry; Biosensing Techniques; Calibration; Cell metabolism and mitochondrial function; Cell Survival; Cells; Cellular oxygen; Characterization and Evaluation of Materials; Chemistry; Chemistry and Materials Science; Comparative analysis; Coproporphyrins - chemistry; Fluorescent Dyes - chemical synthesis; Fluorescent Dyes - chemistry; Food Science; Humans; Infection; Intracellular probe; Investigations; Laboratory Medicine; Luminescent Measurements; Mathematical analysis; Molecular Probes - chemical synthesis; Molecular Probes - chemistry; Molecular Structure; Monitoring/Environmental Analysis; Organoplatinum Compounds - chemical synthesis; Organoplatinum Compounds - chemistry; Original Paper; Oxygen - analysis; Oxygen - metabolism; Phosphorescent oxygen-sensitive probes; Platinum; Pt-porphyrins; Rats; Sensitivity and Specificity; Time Factors; Tumor Cells, Cultured
• ISSN: 1618-2642; 1618-2650
• DOI: 10.1007/s00216-009-3399-z

Several new derivatives of the phosphorescent Pt(II)-coproporphyrin (PtCP) were evaluated with respect to the sensing of intracellular oxygen by phosphorescence quenching. Despite the more favorable molecular charge compared to PtCP, self-loading into mammalian cells was rather inefficient for all the dyes, while cell loading by facilitated transport using transfection reagents produced promising results. The PtCP-NH₂ derivative, which gave best loading efficiency and S/N ratio, was investigated in detail including the optimisation of loading conditions, studies of sub-cellular localization, cytotoxicity, oxygen sensitivity and long-term signal stability. Being spectrally similar to the macromolecular MitoXpress™ probe currently used in this application, the PtCP-NH₂ demonstrated higher loading efficiency and phosphorescent signals, suitability for several problematic cell lines and a slightly increased lifetime scale for the physiological range (0-200 μM O₂). In physiological experiments with different cell types, mitochondrial uncouplers and inhibitors performed on a time-resolved fluorescence plate reader, this probe produced the anticipated profiles of intracellular oxygen concentration and responses to cell stimulation. Therefore, PtCP-NH₂ represents a convenient probe for the experiments and applications in which monitoring of cellular oxygen levels is required.