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Picrosirius red

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Picrosirius Red is a staining reagent used in histological analysis. It selectively binds to collagen fibers, allowing for the visualization and differentiation of various collagen types under polarized light microscopy.

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Lab products found in correlation

Trifluoroacetic acid, by Thermo Fisher Scientific (1 mentions) Iodoacetamide, by Bio-Rad (1 mentions) Acetonitrile, by Thermo Fisher Scientific (1 mentions) Formic acid, by Thermo Fisher Scientific (1 mentions) Chondroitinase abc, by Merck Group (1 mentions) Paraformaldehyde solution, by Thermo Fisher Scientific (1 mentions) Superfrost plus microscope slide, by Thermo Fisher Scientific (1 mentions) Ammonium hydroxide, by Merck Group (1 mentions) Ammonium acetate, by Merck Group (1 mentions) Hyaluronidase, by Merck Group (1 mentions) Dithiothreitol (dtt), by Merck Group (1 mentions) Ammonium bicarbonate, by Merck Group (1 mentions) Mac 3, by BD (1 mentions) Alcian blue, by Thermo Fisher Scientific (1 mentions) Alkaline phosphatase, by Vector Laboratories (1 mentions) Ab222941, by Abcam (1 mentions) Ab210579, by Abcam (1 mentions) C6219, by Merck Group (1 mentions) A5228, by Merck Group (1 mentions) Toluidine blue, by Thermo Fisher Scientific (1 mentions)

5 protocols using picrosirius red

1

Extracellular Matrix Protein Analysis

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Hyaluronidase, chondroitinase ABC, dithiothreitol, ammonium hydroxide, ammonium acetate, and ammonium bicarbonate were obtained from Sigma. Iodoacetamide was obtained from Biorad. Heparinases I, II, and III were obtained from New England Biolabs. Trypsin was obtained from Prozyme. Superfrost Plus microscope slides, picrosirius red, 4% paraformaldehyde solution, 200-proof ethyl alcohol, trifluoroacetic acid, acetonitrile, water, and formic acid were obtained from Fisher Scientific. Xylene and C18 micro-ziptips were obtained from Millipore.
Downs M., Sethi M.K., Raghunathan R., Layne M.D, & Zaia J. (2022). Matrisome changes in Parkinson’s disease. Analytical and bioanalytical chemistry, 414(9), 3005-3015.
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2

Immunohistochemical Analysis of Tissue Samples

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Tissues were fixed with 4% paraformaldehyde at 4 degrees Celsius, dehydrated in 70% ethanol, and embedded in paraffin for sectioning. Using standard immunohistochemical techniques, slides were stained for macrophages (Mac-3, BD Pharmingen, 550292, rat anti-mouse Mac-3 raised against mouse C57Bl/6 peritoneal exudate cells, dilution 1:500), endothelial cells (vWF, Chemicon International, AB7356, rabbit polyclonal antibody raised against human vWF purified from plasma, dilution 1:200 to 1:100), and collagen (picrosirius red, Fisher Scientific). For each sample, 6 high powered fields (HPF) were analyzed using ImageJ software (U.S. National Institutes of Health, Bethesda, MD, USA, http://imagej.nih.gov/ij/).
Nguyen K.P., McGilvray K.C., Puttlitz C.M., Mukhopadhyay S., Chabasse C, & Sarkar R. (2015). Matrix Metalloproteinase 9 (MMP-9) Regulates Vein Wall Biomechanics in Murine Thrombus Resolution. PLoS ONE, 10(9), e0139145.
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3

Tissue Histology and Staining Protocol

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Samples were fixed in 10% Neutralised Buffered Formalin (Sigma) at room temperature overnight, dehydrated in graded alcohols, paraffin embedded and sectioned at 4μm. Tissue slides were stained according to manufacturers’ instructions with Haematoxylin and Eosin (H&E) (Thermo Fisher), Alkaline Phosphatase (Vector Laboratories), Alcian-Blue Periodic Acid Schiff (Sigma). Picrosirius Red (PR), Elastic Van Gieson (EVG) and Alcian Blue (AB) (Thermo Fisher) staining was used to assess retention of collagen, elastin and glycosaminoglycans respectively.
Laween M., Isobel M., Sara C., Weston A.E., Riana G., Lucinda T., Peter F., Michael O., Anna K., Anna B., Laura N., Nikolaos A., Elizabeth H., Julia K., Tedeschi A.M., Pellegata A.F., Susanna E., Alessandro A.P., Lucy C., Nikhil T., Thomas G.M., Simon E., Paola B., De Coppi P, & Li V.S. (2020). Engineering transplantable jejunal mucosal grafts using patient-derived organoids from children with intestinal failure. Nature medicine, 26(10), 1593-1601.
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4

Histological and Biochemical Analysis of Atrial Tissue

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Atria were collected, fixed, and sectioned for histological analysis of alpha smooth muscle actin (αSMA; A5228, Sigma), inflammatory infiltrates (hematoxylin and eosin (H&E), Sigma), collagen (ab210579, Abcam; LS-F5563-1, LifeSpan BioSciences; picrosirius red, Thermo Fisher Scientific), wheat germ agglutinin (Thermo Fisher Scientific), connexin 40 (ab1726, Millipore Sigma), and connexin 43 (C6219, Sigma). Images were quantified using ImageJ 37 (link), 38 (link). Left atria collagen content was confirmed using an assay for hydroxyproline (ab222941, Abcam), an indicator of collagen content/turnover 39 (link), 40 (link).
Homogenized atrial tissue was analyzed for markers associated with RTFI00051 using enzyme-linked immunosorbent assays (ELISAs). These assays included matrix metalloproteinase 2 (MMP2; RTFI00042, AssayGenie), MMP9 (RTFI00080, AssayGenie), tissue inhibitor of metalloproteinases 2 (TIMP2; RTFI00051, AssayGenie), TIMP3 (RTFI01169, AssayGenie), and αSMA (MBS2703516, MyBioSource).
Parent S., Vaka R., St Amant J., Kahn S., Van Remortel S., Bi C., Courtman D., Stewart D.J, & Davis D.R. (2024). Inactivation of the NLRP3 inflammasome mediates exosome-based prevention of atrial fibrillation. Theranostics, 14(2), 608-621.
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5

Decellularization and ECM Characterization

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To confirm decellularization and assess the presence and distribution of glycosaminoglycans (GAGs) and collagen, DNP and native NP tissue samples were fixed, paraffin-embedded, and sectioned at a thickness of 7 μm, prior to staining with either 4′,6-diamidino-2-phenylindole (DAPI), Toluidine blue (0.1%, Sigma), or Picrosirius red (1.3% picric acid and 0.1% direct red 80 (Sigma)), using previously established methods (Shridhar et al., 2019 (link), 2020 (link)). DAPI staining was visualized using an EVOS FL fluorescence microscope (ThermoFisher Scientific Inc.), and Toluidine blue and Picrosirius red staining were visualized using an EVOS XL Core microscope (ThermoFisher Scientific Inc., Burlington, Canada) and a Nikon Optiphot polarizing microscope (Nikon Instruments Inc., Melville, United States), respectively.
Herrera Quijano M.A., Sharma N., Morissette Martin P., Séguin C.A, & Flynn L.E. (2022). Development of 2-D and 3-D culture platforms derived from decellularized nucleus pulposus. Frontiers in Bioengineering and Biotechnology, 10, 937239.
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