A method for enzyme- and immunohistochemical staining of large frozen specimens

Holmbom, Birgitta; Lindström, Mona; Näslund, Ulf; Thornell, Lars‐Eric

doi:10.1007/bf00315739

Cited by 4 publications

(4 citation statements)

References 23 publications

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“…In the samples of experimental myocardial infarction, all the three antibodies against fibrinogen showed distinct intracellular staining in necrotic cardiomyocytes as previously shown [38]. However, in the soleus muscle samples, positive staining for fibrinogen as small dots was seen in some areas in both muscle fibres and extracellular matrix (Fig.…”

Section: Resultssupporting

confidence: 79%

“…Fibrinogen has been previously proved to be a good marker for cardiomyocyte damage after myocardial infarction [38]. However, in the present study, some small intracellular staining for fibrinogen existed not only in post exercise muscles (<5%) but also in controls (around 3.7%).…”

Section: Discussioncontrasting

confidence: 67%

“…Muscle fibre membrane integrity was also evaluated through identification of cellular distribution of plasma proteins fibronectin and fibrinogen. Fibronectin has been previously used as an indicator for membrane injury in muscle fibre [18] and fibrinogen as a sensitive marker for membrane damage in cardiomyocytes after myocardial infarction [38]. Myocardial tissue samples obtained from experimental myocardial infarction in pigs were used as control for specificity of the staining reaction.…”

Section: Methodsmentioning

confidence: 99%

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Re-Evaluation of Sarcolemma Injury and Muscle Swelling in Human Skeletal Muscles after Eccentric Exercise

Liu

Carlsson

et al. 2013

PLoS ONE

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The results regarding the effects of unaccustomed eccentric exercise on muscle tissue are often conflicting and the aetiology of delayed onset muscle soreness (DOMS) induced by eccentric exercise is still unclear. This study aimed to re-evaluate the paradigm of muscular alterations with regard to muscle sarcolemma integrity and fibre swelling in human muscles after voluntary eccentric exercise leading to DOMS. Ten young males performed eccentric exercise by downstairs running. Biopsies from the soleus muscle were obtained from 6 non-exercising controls, 4 exercised subjects within 1 hour and 6 exercised subjects at 2–3 days and 7–8 days after the exercise. Muscle fibre sarcolemma integrity, infiltration of inflammatory cells and changes in fibre size and fibre phenotype composition as well as capillary supply were examined with specific antibodies using enzyme histochemistry and immunohistochemistry. Although all exercised subjects experienced DOMS which peaked between 1.5 to 2.5 days post exercise, no significant sarcolemma injury or inflammation was detected in any post exercise group. The results do not support the prevailing hypothesis that eccentric exercise causes an initial sarcolemma injury which leads to subsequent inflammation after eccentric exercise. The fibre size was 24% larger at 7–8 days than at 2–3 days post exercise (p<0.05). In contrast, the value of capillary number per fibre area tended to decrease from 2–3 days to 7–8 days post exercise (lower in 5 of the 6 subjects at 7–8 days than at 2–3 days; p<0.05). Thus, the increased fibre size at 7–8 days post exercise was interpreted to reflect fibre swelling. Because the fibre swelling did not appear at the time that DOMS peaked (between 1.5 to 2.5 days post exercise), we concluded that fibre swelling in the soleus muscle is not directly associated with the symptom of DOMS.

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Section: Resultssupporting

confidence: 79%

Section: Discussioncontrasting

confidence: 67%

Section: Methodsmentioning

confidence: 99%

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Re-Evaluation of Sarcolemma Injury and Muscle Swelling in Human Skeletal Muscles after Eccentric Exercise

Liu

Carlsson

et al. 2013

PLoS ONE

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“…After perfusion animals were quickly placed into isopentane cooled with dry ice (approximately ¡50°C for 30 s and then stored at ¡70°C until sectioning. Frozen animal was cryosectioned using a heavy-duty cryomicrotome (LKB 2250, LKB, Stockholm, Sweden) into 100 m sagittal sections, which were transferred to the gelatinized glass plates as previously described (Ullberg and Larsson 1981;Kawamoto and Shimizu 1986;Holmbom et al 1991).…”

Section: Animals and Tissue Preparationmentioning

confidence: 99%

Distribution of prolyl oligopeptidase in the mouse whole-body sections and peripheral tissues

Myöhänen

Venäläinen

Garcı́a-Horsman

et al. 2008

Histochem Cell Biol

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Prolyl oligopeptidase (POP) is a serine endopeptidase that hydrolyses proline-containing peptides shorter than 30-mer, including many bioactive peptides. The distribution of POP in the brain has been studied but little is known about the distribution of peripheral POP. We used immunohistochemistry to localize POP in mouse wholebody sections and at the cellular level in peripheral tissues. Furthermore, we used a POP activity assay to reveal the associations between POP protein and its enzymatic activity. The highest POP protein densities were found in brain, kidney, testis and thymus, but in the liver the amounts of POP protein were small. There were remarkable diVerences between the distribution of POP protein and activity. The highest POP activities were found in the liver and testis while kidney had the lowest activity. In peripheral tissues, POP was present in various cell types both in the cytoplasm and nucleus of the cells, in contrast to the brain where no nuclear localization was detected. These Wndings support the proposed role of POP in cell proliferation in peripheral tissues. The dissociation of the distribution of POP protein and its enzymatic activity points to nonhydrolytic functions of POP and to strict endogenous regulation of POP activity.

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Comparison of triphenyltetrazolium chloride (TTC) staining versus detection of fibronectin in experimental myocardial infarction

et al. 1993

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Staining with triphenyltetrazolium chloride (TTC), although controversial, has frequently been used for the delineation of myocardial infarction. This study was performed further to explore the reliability of the TTC method. In 24-h experiments pigs were subjected to closed-chest occlusion of the left anterior descending coronary artery for 30, 60 or 90 min followed by reperfusion with or without superoxide dismutase (SOD) as an adjunct. One TTC-stained slice from each heart was stabilized by microwave irradiation, gelatin-embedded, frozen in hexane chilled with dry ice and cryosectioned. Serial sections were stained with antibodies against fibronectin in order to identify irreversibly injured myocytes and with van Gieson histologically to confirm the necrotic tissue. A close correspondence of the infarct size was found between TTC stained slices and anti-fibronectin stained sections. The infarct size in the van Gieson stained sections also showed good correspondence but the area of infarction tended to be larger. In the experimental group subjected to 30 min ischaemia and with SOD as an adjunct, the estimated infarcted area in the TTC stained slices was significantly smaller than the area estimated from the anti-fibronectin stained sections. In sections viewed in the light microscope an inverse pattern of TTC and anti-fibronectin staining was observed. It was confirmed at the light microscopic level that myocytes containing an abundance of TTC deposits lacked fibronectin whereas myocytes stained with antifibronectin in general lacked TTC staining except for a zone approximately 0.5 mm wide which was located at the intersection between damaged and surviving myocytes where small TTC deposits were present. The width of the stained zone did not differ among the experimental groups. Thus, differences in estimated infarct size by the three methods used reflect problems in correctly delineating the border between living and dead myocardium rather than an interference by SOD on TTC staining.

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A method for enzyme- and immunohistochemical staining of large frozen specimens

Cited by 4 publications

References 23 publications

Re-Evaluation of Sarcolemma Injury and Muscle Swelling in Human Skeletal Muscles after Eccentric Exercise

Re-Evaluation of Sarcolemma Injury and Muscle Swelling in Human Skeletal Muscles after Eccentric Exercise

Distribution of prolyl oligopeptidase in the mouse whole-body sections and peripheral tissues

Comparison of triphenyltetrazolium chloride (TTC) staining versus detection of fibronectin in experimental myocardial infarction

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