Storage-associated artefact in equine muscle biopsy samples.
Abstract: Muscle biopsy is increasingly used in equine veterinary practice for investigating exertional, inflammatory or immune mediated myopathies and unexplained muscle atrophy. Although formalin-fixed samples are often used, for complete evaluation, fresh-frozen tissue is required. Freezing muscle in veterinary practice is impractical: samples sent to specialist laboratories for processing are therefore susceptible to delays, potentially leading to artefact and compromising histological interpretation. Objective: Altered temperature, duration and hydration status influence the severity of storage-induced artefact in equine muscle. Methods: Skeletal muscle obtained immediately post euthanasia was divided into 6 independent samples from each of 8 horses. One sample per horse was frozen immediately in isopentane precooled in liquid nitrogen. Additional samples were stored in conditions designed to mimic possible situations encountered in practice, including increased storage times, temperature and hydration status. Following storage, stored samples were frozen as before. Cryosections were stained using haematoxylin and eosin and ranked for artefact on 2 occasions by 2 blinded observers. The best samples were processed subsequently with a panel of routine stains and immunolabelled for collagen V to enable the measurement of minimum fibre diameters. Results: Both prolonged storage and increased hydration resulted in more storage-associated artefact. Samples stored for 24 h chilled on dry gauze were ranked higher than those stored on damp gauze; however, a panel of routinely-used histochemical staining techniques was unaffected by chilled 24 h storage. There was no significant effect of storage on mean fibre diameter; however, both chilled dry and damp storage for 24 h caused a significant increase in fibre-size variability. Conclusions: Caution should be exercised when interpreting fibre size profiles in shipped samples. Equine muscle biopsy samples are optimally shipped in dry gauze, sealed in plastic containers and shipped on ice packs to be processed within 24 h and can thus be interpreted by the receiving laboratory with minimal artefact.
Publication Date: 2009-03-24 PubMed ID: 19301587DOI: 10.2746/042516408x330374Google Scholar: Lookup
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- Journal Article
Summary
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This research scrutinizes how the severity of storage-induced artifacts in horse muscle samples is impacted by alterations in temperature, storage duration, and hydration status. The study advocates careful shipment of these samples in dry gauze, sealed in plastic, and transported on ice packs within 24 hours to minimize artifacts.
Objective and Methodology
- The central goal of this research was to understand how storage conditions, specifically varying temperature, storage time and hydration levels could affect the quality of equine muscle tissue biopsy samples. These factors are observed to influence the occurrence of storage-induced artifacts, which can potentially compromise the histological interpretation of the samples.
- The experiment consisted of obtaining skeletal muscle samples from 8 horses immediately post euthanasia. Six independent samples were taken from each horse. While one sample was frozen immediately using isopentane precooled in liquid nitrogen, the rest were stored under different conditions to replicate possible situations encountered in practice.
- After storage, these samples were again frozen under similar conditions and subsequently stained using haematoxylin and eosin. They were then evaluated for artifacts by two different blinded observers.
- The best quality samples were further processed with a range of routine stains and marked for collagen V to facilitate measurement of minimum fiber diameters.
Results
- The main findings of the study highlight that prolonged storage and increased hydration led to more storage-associated artifacts in the muscle samples. Samples stored for 24 hours in dry gauze at low temperature were found to be better in quality as compared to those placed on damp gauze.
- Notably, the study also found the histochemical staining techniques typically used in practice were not impacted by 24 hours of chilled storage.
- Interestingly, the average fiber diameter was not significantly affected by storage. However, storage in either dry or damp conditions for 24 hours did increase the variability in fiber sizes significantly. This suggests a shift in the distribution of fiber sizes rather than a generalized change being induced due to storage.
Conclusions
- Care needs to be exercised when interpreting fiber size profiles in shipped samples. It is possible that storage-associated artifacts might affect our understanding of these attributes.
- The research suggests that equine muscle biopsy samples are ideally shipped in dry gauze, sealed in plastic containers, and transported on ice packs. If these conditions are maintained so that the samples can be processed within 24 hours, the likelihood of encountering storage-induced artifacts can be minimized. This, in turn, enables the receiving laboratory to interpret the samples with minimal artifacts.
Cite This Article
APA
Stanley RL, Maile C, Piercy RJ.
(2009).
Storage-associated artefact in equine muscle biopsy samples.
Equine Vet J, 41(1), 82-86.
https://doi.org/10.2746/042516408x330374 Publication
Researcher Affiliations
- Comparative Neuromuscular Diseases Laboratory, Department of Veterinary Clinical Sciences, The Royal Veterinary College, Hawkshead Lane, North Mymms, Haffield, Hertfordshire AL9 7TA, UK.
MeSH Terms
- Animals
- Artifacts
- Biopsy / methods
- Biopsy / standards
- Biopsy / veterinary
- Cryopreservation / veterinary
- Horse Diseases / diagnosis
- Horse Diseases / pathology
- Horses
- Immunohistochemistry / veterinary
- Muscle Fibers, Skeletal / pathology
- Muscular Atrophy / diagnosis
- Muscular Atrophy / pathology
- Muscular Atrophy / veterinary
- Muscular Diseases / diagnosis
- Muscular Diseases / pathology
- Muscular Diseases / veterinary
- Observer Variation
- Time Factors
- Tissue and Organ Harvesting / methods
- Tissue and Organ Harvesting / standards
- Tissue and Organ Harvesting / veterinary
Citations
This article has been cited 2 times.- Bookbinder L, Finno CJ, Firshman AM, Katzman SA, Burns E, Peterson J, Dahlgren A, Ming-Whitfield B, Glessner S, Borer-Matsui A, Valberg SJ. Impact of alpha-tocopherol deficiency and supplementation on sacrocaudalis and gluteal muscle fiber histopathology and morphology in horses. J Vet Intern Med 2019 Nov;33(6):2770-2779.
- Moore CD, Fahlman A, Crocker DE, Robbins KA, Trumble SJ. The degradation of proteins in pinniped skeletal muscle: viability of post-mortem tissue in physiological research. Conserv Physiol 2015;3(1):cov019.
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