Detecting microcalcifications in atherosclerotic plaques by a simple trichromic staining method for epoxy embedded carotid endarterectomies.
Atherosclerotic plaques have a high probability of undergoing rapid progression to stenosis, becoming responsible of acute coronary syndrome or stroke. Microcalcifications may act as enhancers of atherosclerotic plaque vulnerability. Considering that calcifications with a diameter smaller than 10 mm in paraffin embedded tissue are rather difficult to detect, our aim was to analyze microcalcifications on semithin sections from epoxy resin embedded samples of carotid endarterectomies using an original trichromic stain (methylene blue--azur B--basic fuchsine--alizarin red). We have compared samples stained either with our method, methylene blue-azur B alone or with Von Kossa staining, and methylene blue-azur B -basic fuchsine alone or with Von Kossa staining. Our method resulted to be simple and fast (ca. 2 min), it gives a sharp general contrast for all structures and allows to easy identify collagen and elastin. In addition, gray-green colour associated to intracellular lipid droplets evidences foam cells, which are particularly abundant in endarterectomies samples. Mast cells and their metachromatic granules are also well recognized. Calcifications over 0,5 mm are clearly recognizable. In conclusion, microcalcifications are clearly distinguished from the extracellular matrix in spite of their reduced dimensions. Methylene blue--azur B--basic fuchsine--alizarin red method is easy to use, reproducible, and is particularly suitable for the identification of microcalcifications in the morphological analysis of atherosclerotic plaques.
Relucenti
M
M
Laboratory of Electron Microscopy Pietro M. Motta, Department of Human Anatomic, Histologic, Forensic and Locomotor Apparatus Sciences, Sapienza University of Rome, via Alfonso Borelli 50, Rome, Italy. michela.relucenti@uniroma1.it
Heyn
R
R
Petruzziello
L
L
Pugliese
G
G
Taurino
M
M
Familiari
G
G
D000368:Aged; D002114:Calcinosis / Q000175:diagnosis*; D016894:Endarterectomy, Carotid; D004853:Epoxy Resins; D005260:Female; D006801:Humans; D008297:Male; D046529:Microscopy, Electron, Transmission; D008875:Middle Aged; D058226:Plaque, Atherosclerotic / Q000175:diagnosis* / Q000648:ultrastructure; D013194:Staining and Labeling / Q000379:methods*
D004853:Epoxy Resins
Stain Technol. 1973 Sep;48(5):223-7
4126687
Z Kardiol. 2001;90 Suppl 3:68-80
11374037
Circulation. 2003 Oct 7;108(14):1664-72
14530185
Cerebrovasc Dis. 2004;18(2):117-23
15218276
Anal Chem. 2008 Jan 1;80(1):55-61
18027908
Stain Technol. 1974 Jan;49(1):9-14
4131626
J Histochem Cytochem. 2001 Mar;49(3):397-406
11181742
J Histochem Cytochem. 1958 Jan;6(1):22-42
13514045
Stroke. 2005 May;36(5):1091-7
15774817
Calcif Tissue Int. 2003 May;72(5):537-47
12724828
J Biomech. 2008;41(5):1111-8
18258240
Proc Natl Acad Sci U S A. 2006 Oct 3;103(40):14678-83
17003118
Arch Histol Cytol. 2000 Jul;63(3):271-84
10989939
J Microsc. 2008 Dec;232(3):504-7
19094029
Prog Histochem Cytochem. 1985;16(2):1-84
2417278
Stroke. 2008 Jun;39(6):1722-9
18403733
Biotech Histochem. 2005 Sep-Dec;80(5-6):207-10
16720521
J Pathol. 1998 May;185(1):10-7
9713354
J Clin Invest. 1994 Oct;94(4):1597-604
7929835
J Histochem Cytochem. 2003 Feb;51(2):199-204
12533528
Appl Immunohistochem Mol Morphol. 2005 Sep;13(3):292-4
16082259
Mol Cell Biomech. 2008 Mar;5(1):37-47
18524245
Micron. 2004;35(4):287-96
15003616
Histochemistry. 1994 Jan;101(1):51-5
7517926
Stain Technol. 1986 Mar;61(2):89-92
2424151