Nondestructive Online In Vitro Monitoring of Pre-Osteoblast Cell Proliferation Within Microporous Polymer Scaffolds

We present a system for the online, in vitro, nondestructive monitoring of tissue growth within microporous polymer scaffolds. The system is based on measuring the admittance of the sample over a frequency range of 10-200 MHz using an open-ended coaxial probe and impedance analyzer. The sample admittance is related to the sample complex permittivity (CP) by a quasi-static model of the probe's aperture admittance. A modified effective medium approximation is then used to relate the CP to the cell volume fraction. The change of cell volume fraction is used as a measure of tissue growth inside the scaffold. The system detected relative cell concentration differences between microporous polymer scaffolds seeded with 0.4, 0.45, 0.5, and 0.6 x 106 pre-osteoblast cells. In addition, the pre-osteoblast proliferation within 56 scaffolds over 14 days was recorded by the system and a concurrent DNA assay. Both techniques produced cell proliferation curves that corresponded to those found in literature. Thus, our data confirmed that the new system can assess relative cell concentration differences in microporous scaffolds enabling online nondestructive tissue growth monitoring.

[1]  U von Stockar,et al.  On-line determination of animal cell concentration in two industrial high-density culture processes by dielectric spectroscopy. , 2002, Biotechnology and bioengineering.

[2]  I. Woodhead,et al.  An Open-ended Coaxial Probe for Broad-band Permittivity Measurement of Agricultural Products , 1999 .

[3]  U A Stock,et al.  Tissue engineering: current state and prospects. , 2001, Annual review of medicine.

[4]  D. Hutmacher,et al.  Scaffolds in tissue engineering bone and cartilage. , 2000, Biomaterials.

[5]  Omowunmi A. Sadik,et al.  Impedance Spectroscopy: A Powerful Tool for Rapid Biomolecular Screening and Cell Culture Monitoring , 2005 .

[6]  Dietmar W Hutmacher,et al.  Current strategies for cell delivery in cartilage and bone regeneration. , 2004, Current opinion in biotechnology.

[7]  Christopher L. Davey,et al.  The dielectric properties of biological cells at radiofrequencies : Applications in biotechnology , 1999 .

[8]  Dietmar W Hutmacher,et al.  The challenge to measure cell proliferation in two and three dimensions. , 2005, Tissue engineering.

[9]  M. Tabrizian,et al.  Complex permittivity measurement as a new noninvasive tool for monitoring in vitro tissue engineering and cell signature through the detection of cell proliferation, differentiation, and pretissue formation , 2004, IEEE Transactions on NanoBioscience.

[10]  Devendra K. Misra,et al.  A study on the coaxial aperture electromagnetic sensor and its application in material characterization , 1993 .

[11]  C. Cametti,et al.  Dielectric spectroscopy of erythrocyte cell suspensions. A comparison between Looyenga and Maxwell-Wagner-Hanai effective medium theory formulations , 2002 .

[12]  Daniël De Zutter,et al.  Design rules for an experimental setup using an open-ended coaxial probe based on theoretical modelling , 1994 .

[13]  M. A. Stuchly,et al.  Equivalent circuit of an open-ended coaxial line in a lossy dielectric , 1982, IEEE Transactions on Instrumentation and Measurement.

[14]  Christopher L. Davey,et al.  The influence of electrode polarisation on dielectric spectra, with special reference to capacitive biomass measurements I. Quantifying the effects on electrode polarisation of factors likely to occur during fermentations , 1998 .

[15]  L. Griffith,et al.  Tissue Engineering--Current Challenges and Expanding Opportunities , 2002, Science.

[16]  Monitoring of stem cell proliferation and differentiation using a permittivity-responsive biointerface , 2003 .

[17]  D. K. Misra,et al.  A Quasi-Static Analysis of Open-Ended Coaxial Lines (Short Paper) , 1987 .

[18]  Koji Asami,et al.  Characterization of biological cells by dielectric spectroscopy , 2002 .

[19]  K. Zhao,et al.  Dielectric spectroscopy of Anabaena 7120 protoplast suspensions. , 2006, Bioelectrochemistry.

[20]  S. Gawad,et al.  Single cell dielectric spectroscopy , 2007 .

[21]  M J Yaszemski,et al.  Three-dimensional culture of rat calvarial osteoblasts in porous biodegradable polymers. , 1998, Biomaterials.

[22]  P. Bianco,et al.  Stem cells in tissue engineering , 2001, Nature.

[23]  D. Hungerford,et al.  Chitosan supports the expression of extracellular matrix proteins in human osteoblasts and chondrocytes. , 2000, Journal of biomedical materials research.

[24]  R. D. Pollard,et al.  An improved technique for permittivity measurements using a coaxial probe , 1997 .

[26]  J Marler,et al.  Transplantation of cells in matrices for tissue regeneration. , 1998, Advanced drug delivery reviews.

[27]  Teodor Veres,et al.  Towards on‐line monitoring of cell growth in microporous scaffolds: Utilization and interpretation of complex permittivity measurements , 2003, Biotechnology and bioengineering.

[28]  P. Żółtowski,et al.  On the electrical capacitance of interfaces exhibiting constant phase element behaviour , 1998 .

[29]  Devendra K. Misra,et al.  REVIEW ARTICLE: The co-axial aperture electromagnetic sensor and its application in material characterization , 1997 .

[30]  S. Madihally,et al.  Porous chitosan scaffolds for tissue engineering. , 1999, Biomaterials.

[31]  John Rollo,et al.  Biomaterials and scaffold design: key to tissue‐engineering cartilage , 2007, Biotechnology and applied biochemistry.

[32]  Dietmar W. Hutmacher,et al.  Biodegradable polymers applied in tissue engineering research: a review , 2007 .

[33]  J. Dibeneditto,et al.  Frequency dependence of 50-Ω coaxial open-circuit reflection standard , 1981, IEEE Transactions on Instrumentation and Measurement.

[34]  B. Colpitts.  Temperature sensitivity of coaxial probe complex permittivity measurements: experimental approach , 1993 .