Effects of freezing and short‐term fixation on muscle mass, volume, and density

Abstract

Preventing postmortem deterioration of soft-tissues is an important requisite of anatomical research. In order to provide corrections for potential myological distortions, this study quantifies the acute effects of freezing, formalin fixation and ethanol storage using muscles from (n = 46) rabbits (Oryctolagus cuniculus). Bilateral dissections of specific muscles were performed and each side was assigned to a different preparation group (fresh, formalin fixation only, fixation followed by short duration ethanol storage, and freezing once or twice). We demonstrate that short-term freezing at À20C and thawing have no significant effect on muscle mass, volume, and density while short-term formalin fixation and ethanol storage significantly reduces mass and volume (density remains relatively constant.) Although freezing may have less of an effect on the gross morphometric characteristics of the musculature than ethanol storage, slow freezing damages muscle microanatomy, and therefore, faster freezing and other modes of preservation such as formalin fixation and ethanol storage may be preferable. Based on our results, we derived the following correction factors for each preparation: the mass of specimens stored in 70% ethanol should be multiplied by 1.69 to approximate fresh muscle mass, and specimens fixed in 10% formalin multiplied by 1.32. Although not significant, specimens frozen-once were slightly less massive and could be multiplied by 1.03 (frozen-twice Â1.09). The volumetric corrections are: ethanol 1.64; 10% formalin 1.32; frozen-once 1.03; frozen-twice 1.10. While the density of ethanol preserved specimens is slightly less than that of fresh ones (correction: 1.03), those preserved in formalin and frozen maintain nearly the same density.

Three-dimensional polygonal muscle modelling and line of action estimation in living and extinct taxa

“…The volumetric muscle models created using the iterative polygonal modelling approach (either surface-based or tomography-guided) can be evaluated by the criterion that simplifications and/or predictions should quantitatively fall within the range of physical measurements or follow observed patterns 21 , 29 , 33 . The 3D muscle masses of both the diceCT segmented and the reconstructed muscles using the iterative polygonal modelling approach were calculated from their volumes with an assumed homogeneous density of 1060 kg/m 3 3 , 102 , 126 , 127 . These calculated muscle masses of the Nile crocodile were then compared to the median of the body-mass-normalised measurements of the respective muscles of other Nile crocodile specimens (Fig.…”

Scaling of fibre area and fibre glycogen concentration in the hindlimb musculature of monitor lizards: implications for locomotor performance with increasing body size

“…All fCSA values were multiplied by a conversion factor of 1.63212 to account for the reduction in fibre cross-sectional area that occurs as a result of 1 month of ethanol storage according to published decay equations (Leonard et al, 2021a,b). Storage in ethanol for 1 month after fixation in formalin causes a decrease in muscle mass of 41% compared with fresh tissue (Leonard et al, 2021a) and reductions in fCSA relate to reductions in muscle mass over longer periods by a reduced major axis regression [relative fCSA=(relative muscle mass×10,113.5)+5.9272; R 2 =0.9648] (Leonard et al, 2021b) such that fCSA can be expected to decrease by 61.27%. To control for differences in specimen preparation, fCSA was calculated among all samples, and also among a subset of the data where specimens were never frozen prior to analysis.…”

Heat Emitting Damage in Skin: A Thermal Pathway for Mechanical Algesia

“…This being stated, we chose skin samples where the dehairing damage was the less pronounced. The second potentially altering process is freezing, which can cause micro-damages in soft biological tissues (e.g., Leonard et al, 2021). The skin samples were indeed bought frozen and were unfrozen in a ambient temperature water bath during half an hour before running the rupture tests.…”