The effect of preparation technique on the optical parameters of biological tissue

Abstract

The absorption coefficient μa, the scattering coefficient μs, and the scattering anisotropy factor g of porcine liver were studied in vitro using the integrating sphere technique and inverse Monte Carlo simulation in the wavelength range 450 to 700 nm. A reference preparation technique was developed using a dermatome providing specimens of 200 to 800 μm thickness without pre-freezing the tissue. The optical parameters as measured applying the reference preparation were compared to those measured after cryo-homogenisation. We found significant deviations of the scattering coefficient and the anisotropy factor which were compensated when the reduced scattering coefficient μs ′ was calculated. We also compared the effects of freezing reference specimens at -20 °C and at 77 K without homogenisation. For both freezing protocols noticeable deviations were found in all three optical parameters as well as in μs ′. The impact of tissue storage at 4 °C was measured in the range 4 to 48 h post mortem and showed a clear reduction of μa and a significant increase of μs even after 24 h of storage. Short-time storage of the specimens in saline solution reduced all three optical parameters significantly. In conclusion, the tissue preparation must be controlled in order to provide in vitro optical parameters that sufficiently mimic the in vivo situation.