Four weeks after AntCaNtide (50 μg/kg), tat-CN17β (50 μg/kg) or NaCl 0.9%, intra-cardiac injection, the hearts were immersion fixed in 10% buffered paraformaldehyde. The tissues were embedded in paraffin, cut at 5 μm, and processed. For Masson trichrome staining of collagen fibers, slides were stained with Weigert Hematoxylin (Sigma-Aldrich, St. Louis, MO) for 10 min., rinsed in PBS (Invitrogen) and then stained with Biebrich scarlet-acid fuchsine (Sigma-Aldrich St. Louis, MO) for 5 min. Slides were rinsed in PBS and stained with phosphomolybdic/phosphotungstic acid solution (Sigma-Aldrich St. Louis, MO) for 5 min. then stained with light green (Sigma-Aldrich) for 5 min. Slides were rinsed in distilled water, dehydrated with 95% and absolute alcohol and a coverslip was placed. For the analysis of cardiomyocytes size, Masson trichrome staining sections were used [37 (link)]. The areas (μm2) of ~100 cardiac myocytes per heart were measured with the public domain Java image processing program Image by an independent operator blind to the study protocol (DS) [44 (link)].
Biebrich scarlet acid fuchsine
Manufactured by Merck Group
Biebrich scarlet-acid fuchsine is a laboratory dye used for staining and visualizing biological samples. It is a mixture of two synthetic dyes, Biebrich scarlet and acid fuchsine, which can be used together or separately for various staining applications.
Automatically generated - may contain errors
Lab products found in correlation
Phosphomolybdic phosphotungstic acid solution, by Merck Group (2 mentions)
Light green, by Merck Group (1 mentions)
Weigert hematoxylin, by Merck Group (1 mentions)
Phosphate buffered saline (pbs), by Thermo Fisher Scientific (1 mentions)
Aniline solution, by Merck Group (1 mentions)
Polylysine slides, by Thermo Fisher Scientific (1 mentions)
Aperio, by Leica Biosystems (1 mentions)
Neutral buffered formalin, by Merck Group (1 mentions)
Prussian blue, by Merck Group (1 mentions)
Mayer s hematoxylin, by Merck Group (1 mentions)
Weigerts hematoxylin, by Carl Roth (1 mentions)
3 3 diaminobenzidine (dab), by Merck Group (1 mentions)
Entellan, by Merck Group (1 mentions)
Acetic acid, by Carl Roth (1 mentions)
2 protocols using biebrich scarlet acid fuchsine
1
Masson Trichrome Analysis of Cardiomyocyte Size
2
Histological Staining Techniques for Tissue Analysis
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Tissues were fixed for 24 h in 10% neutral buffered formalin (Sigma), dehydrated in ethanol gradients, paraffin embedded and sectioned at 3 μm on polylysine slides (Thermo scientific). Slides were rehydrated with decreasing percentage of ethanol solutions and subjected to stains.
For iron staining, slides were quenched with 3% H2O2 and treated for 20 min with Prussian Blue (Sigma Aldrich) followed by a 20 min incubation with 3,3′-diaminobenzidine (DAB) (Sigma Aldrich). Meyer's hematoxylin was used as counterstain.
For Masson's trichrome staining, slides were treated overnight with the Bourin's mordent solution (Roth). After washing, slides were treated for 5 min with Weigert's hematoxylin (Roth), 5 min with Biebrich Scarlet-Acid Fuchsine (Sigma Aldrich), 5 min with Phosphotungstic/Phosphomolybdic Acid Solution (Sigma Aldrich), 5 min with Aniline Solution (Sigma Aldrich) and 2 min with 1% acetic acid (Carl Roth).
For hematoxylin and eosin staining, slides were treated for 6 min with Mayer's Hematoxylin (Sigma Aldrich), rinsed in phosphate buffered saline, and incubated for 2 min in Eosin in acetic acid (Sigma Aldrich).
After the stains, slides were dehydrated with ethanol gradients and mounted with Entellan (Sigma Aldrich). Digital images were acquired up to 40 × magnification with an automated slide scanner (Aperio - Leica biosystems)
For iron staining, slides were quenched with 3% H2O2 and treated for 20 min with Prussian Blue (Sigma Aldrich) followed by a 20 min incubation with 3,3′-diaminobenzidine (DAB) (Sigma Aldrich). Meyer's hematoxylin was used as counterstain.
For Masson's trichrome staining, slides were treated overnight with the Bourin's mordent solution (Roth). After washing, slides were treated for 5 min with Weigert's hematoxylin (Roth), 5 min with Biebrich Scarlet-Acid Fuchsine (Sigma Aldrich), 5 min with Phosphotungstic/Phosphomolybdic Acid Solution (Sigma Aldrich), 5 min with Aniline Solution (Sigma Aldrich) and 2 min with 1% acetic acid (Carl Roth).
For hematoxylin and eosin staining, slides were treated for 6 min with Mayer's Hematoxylin (Sigma Aldrich), rinsed in phosphate buffered saline, and incubated for 2 min in Eosin in acetic acid (Sigma Aldrich).
After the stains, slides were dehydrated with ethanol gradients and mounted with Entellan (Sigma Aldrich). Digital images were acquired up to 40 × magnification with an automated slide scanner (Aperio - Leica biosystems)
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