Optical waveguide lightmode spectroscopy (OWLS) to monitor cell proliferation quantitatively.
The use of microscopic observations used for in situ monitoring of cell proliferation in the production of epidermal autografts is not satisfactory. In particular, the identification of the projected cell area from microscopic pictures by image analysis (IA) depends on intensity edges and level of contrasts and is thus limited to subconfluent cultures. Some of these problems can be solved by using optical waveguide lightmode spectroscopy (OWLS), which measures the effective refractive index of a thin layer above an Si(Ti)O(2) waveguide surface. In this study the use of OWLS to monitor cell adhesion, spreading, and growth was studied. The sensitivity of the method was investigated by using three different cell lines, two fibroblasts and one hepatoma cell line. Cell proliferation of two strains of fibroblasts and hepatoma cells was monitored up to 2 days with the OWLS. In parallel, cell density was determined at different time points microscopically using an additional window in the measuring chamber. The cell density of fully spread cells ( approximately 4 h after attachment) was found to be proportional to the OWLS signal. In long-term cultures the influence of the cell density from single cells to confluent cell cultures upon the OWLS signal was investigated. The exponentially growing number of hepatoma resulted in a linear increase of the sensor signal. Due to this and to the fact that the proliferating cells exhibit contact inhibition, it was concluded that the cell contact area must decrease exponentially. The results show the strength of OWLS for monitoring the adhesion and proliferation of anchorage-dependent cells in applications where an on-line indicator of the total biomass is needed. Additionally, OWLS provides metabolic information through detection of the cell mass in close contact with the waveguide.
Hug
T S
TS
Gnothis SA, PSE-B EPFL, Swiss Federal Institute of Technology (ETH), 1015 Lausanne, Switzerland. thomassilvan@bluemail.ch
Prenosil
J E
JE
Maier
P
P
Morbidelli
M
M
D016475:3T3 Cells / Q000166:cytology / Q000502:physiology; D000818:Animals; D019149:Bioreactors; D015374:Biosensing Techniques / Q000295:instrumentation / Q000379:methods; D006528:Carcinoma, Hepatocellular / Q000473:pathology* / Q000503:physiopathology; D002448:Cell Adhesion / Q000502:physiology; D002452:Cell Count / Q000295:instrumentation / Q000379:methods*; D018929:Cell Culture Techniques / Q000379:methods; D002455:Cell Division; D002478:Cells, Cultured; D003470:Culture Media / Q000032:analysis; D005347:Fibroblasts / Q000166:cytology* / Q000378:metabolism; D006801:Humans; D051379:Mice; D055095:Optics and Photonics / Q000295:instrumentation; D012031:Refractometry / Q000295:instrumentation / Q000379:methods; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D013045:Species Specificity; D013057:Spectrum Analysis / Q000295:instrumentation / Q000379:methods*
D003470:Culture Media