Abstract
This study is aimed at characterizing soft tissue slices using a vibratome. In particular, the effect of two sectioning parameters (i.e., step size and sectioning speed) on resultant slice thickness was investigated for fresh porcine liver as well as for paraformaldehyde-fixed (PFA-fixed) and fresh murine brain. A simple framework for embedding, sectioning and imaging the slices was established to derive their thickness, which was evaluated through a purposely developed graphical user interface. Sectioning speed and step size had little effect on the thickness of fresh liver slices. Conversely, the thickness of PFA-fixed murine brain slices was found to be dependent on the step size, but not on the sectioning speed. In view of these results, fresh brain tissue was sliced varying the step size only, which was found to have a significant effect on resultant slice thickness. Although precision-cut slices (i.e., with regular thickness) were obtained for all the tissues, slice accuracy (defined as the match between the nominal step size chosen and the actual slice thickness obtained) was found to increase with tissue stiffness from fresh liver to PFA-fixed brain. This quantitative investigation can be very helpful for establishing the most suitable slicing setup for a given tissue.
Highlights
Vibrating blade microtomes or vibratomes are commonly used for obtaining precision-cut slices from soft fresh tissues
The two-way ANOVA analysis showed that both the step size and sectioning speed have a significant effect on the resultant thickness of fresh liver slices, 4 of the 6 step size-sectioning speed combinations investigated ( 200-0.1, 200-0.4, 400-0.1 and 400-0.2 μm-mm/s) yielded similar slice thicknesses, with an average value of 540 ± 91 μm (Fig. 3A)
Precision-cut tissue slices from fresh porcine liver as well as from PFA-fixed and fresh murine brain were characterized in their thickness through a semi-automated slice analysis Graphical User Interface (GUI) implemented in Matlab⃝R
Summary
Vibrating blade microtomes or vibratomes are commonly used for obtaining precision-cut slices from soft fresh tissues. Precision-cut liver slices are powerful tools for the in vitro study of pharmacological metabolism, toxicology and efficacy of novel substances under standardized conditions (Van de Bovenkamp et al, 2005; Van de Bovenkamp et al, 2006; Van de Bovenkamp et al, 2007; Karim et al, 2013; Eide et al, 2014). They have been used extensively for rank-ordering the toxicity of chemicals and examining the mechanisms of liver injury as. On the other hand, are attractive for the evaluation of different morphometric features such as the total extent of dendrites and the number of branching points, as well as for 3D tissue reconstruction and analysis of neurons (Jin et al, 2003; Billeci et al, 2013; Golovyashkina et al, 2014)
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