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Impedance Spectroscopy for Toxicity Testing Martin Dominguez Cronus Technologies Ltd.

Published byMarlene Wells Modified over 9 years ago

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Presentation on theme: "Impedance Spectroscopy for Toxicity Testing Martin Dominguez Cronus Technologies Ltd."— Presentation transcript:

1 Impedance Spectroscopy for Toxicity Testing Martin Dominguez Cronus Technologies Ltd.

2 Barrier Forming Tissue Epithelial and endothelial cell layers form selectively permeable barriers

3 The Concept Growing cell layers on permeable membranes

4 The Concept Measuring the electrical impedance of epithelial and endothelial cell layers under physiological conditions

5 Understanding Impedance Spectra Contributions stemming from CPE and R MED

6 Understanding Impedance Spectra The formation of a plateau at mid-range frequencies is attributable to the cell layer’s resistance TER and capacitance C CL

7 Understanding Impedance Spectra An increase in the resistance TER shifts the plateau upwards

8 Understanding Impedance Spectra An increase in the capacitance C CL narrows the plateau

9 Time-Lapse Impedance Spectroscopy

10 The Setup

11

12 cellZscope Automated cell monitoring device

13 Monitoring of MDCK-II Cells Formation of a differentiated cell monolayer with established cell-cell junctions

14 cellZscope vs. „Chopstick“ Electrodes Comparing TER values measured on MDCK-II cell layers

15 Long Term Monitoring of Caco-2 Cells Comparing growth dynamics of different cell lines

16 EDTA Treatment of MDCK-II Cells Effects on the tight junction network

17 MBCD Treatment of MDCK-II Cells Dose dependent response in TER and C cl to cholesterol depletion

18 Testing the Toxicity of Nanoparticles Exposure of MDCK cells to different types of particles + Triton X100 Control Carbon Black * ZrO 2 * ZnO - NP * + Particles * 10 µg/cm² 0 20 40 60 80 100 120 140 160 180 0510152025 Time (h) TER (%) Data courtesy of A. Kroll, J. Schnekenburger, University of Münster, Germany

19 Testing the Toxicity of Nanoparticles Exposure of NRK52E cells to ZnO Control ZnO * ZnO NP * * 10 µg/cm² 0 20 40 60 80 100 120 0510152025 TER (%) + ZnO+ Triton X100 Time (h) Data courtesy of A. Kroll, J. Schnekenburger, University of Münster, Germany

20 cellZscope Automated cell monitoring device

21 Compatibility with Standard Cell Culture Inserts Manufacturer Part No.MembraneInsert 24-Well Size12-Well Size6-Well Size MaterialPore SizePore DensityOpticsType Corning 341334013412PC0.4 µm1.0×10 8 /cm 2 translucenthanging 347034603450PET0.4 µm4.0×10 6 /cm 2 transparenthanging 341534023414PC3.0 µm2.0×10 6 /cm 2 translucenthanging 347234623452PET3.0 µm2.0×10 6 /cm 2 transparenthanging Greiner Bio-One 662 640665 640657 640PET0.4 µm1.0×10 8 /cm 2 translucenthanging 662 641665 641657 641PET0.4 µm2.0×10 6 /cm 2 transparenthanging 662 610665 610657 610PET1.0 µm2.0×10 6 /cm 2 transparenthanging 662 631665 631657 631PET3.0 µm2.0×10 6 /cm 2 translucenthanging 662 630665 630657 630PET3.0 µm6.0×10 5 /cm 2 transparenthanging BD Biosciences 353495353494353090PET0.4 µm1.6×10 8 /cm 2 translucenthanging 353095353180353493PET0.4 µm1.6×10 6 /cm 2 transparenthanging 353104353103353102PET1.0 µm1.6×10 6 /cm 2 transparenthanging 353492353292353092PET3.0 µm2.0×10 6 /cm 2 translucenthanging 353096353180353091PET3.0 µm8.0×10 5 /cm 2 transparenthanging Millipore PIHT 12R 48PIHT 15R 48PIHT 30R 48PET0.4 µm1.0×10 8 /cm 2 translucenthanging PIRP 12R 48PIRP 15R 48PIRP 30R 48PET1.0 µm2.0×10 6 /cm 2 transparenthanging PISP 12R 48PISP 15R 48PISP 30R 48PET3.0 µm2.0×10 6 /cm 2 translucenthanging Nunc 140620140652140660PC0.4 µm8.5×10 7 /cm 2 translucentstanding 140627140654140663PC3.0 µm1.7×10 6 /cm 2 translucentstanding

22 Software for Data Acquisition and Analysis

23 Effects of Hydrocortisone on Endothelial Cells adapted from Wegener et al., BioTechniques 37, 590 (2004) Transepithelial electrical resistance (TER) of primary cultured endothelial cells derived from porcine brain microvessels incubated in serum-free medium supplemented with and without hydrocortisone

24 Effects of Serum-free Media on Epithelial Cells adapted from Wegener et al., BioTechniques 37, 590 (2004) Capacitance (C cl ) of primary cultured epithelial cells derived from porcine choroid plexus incubated in serum-free and serum-containing medium

25 Inflammatory Response to TNF-   Transepithelial electrical resistance (TER) of porcine brain capillary endothelial cells

26 Response to TNF-   Suppressed by Hydrocortisone Transepithelial electrical resistance (TER) of porcine brain capillary endothelial cells

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