Combined orcein and martius scarlet blue (OMSB) staining for qualitative and quantitative analyses of atherosclerotic plaques in brachiocephalic arteries in apoE/LDLR-/-mice.
Numerous cellular and extracellular components should be analyzed in sections of atherosclerotic plaques to assess atherosclerosis progression and vulnerability. Here, we combined orcein (O) staining for elastic fibers and martius scarlet blue (MSB) polychrome to visualize various morphological contents of plaque in brachiocephalic arteries (BCA) of apoE/LDLR-/-mice. Elastic fibers (including broken elastic laminae and 'buried' fibrous caps) were stained purple and they could be easily distinguished from collagen fibers (blue). Orcein allowed clear identification of even the finest elastic fibers. Erythrocytes were stained yellow and they could easily be discerned from mature fibrin (red). Old fibrin tends to acquire blue color. The method of OMSB staining is simple, takes less than 1 h to perform and can be adapted to automatic stainers. Most importantly, the color separation is good enough to allow digital automatic segmentation of specific components in tissue section and quantitative analysis of the plaque constituents. OMSB was used to compare atherosclerotic plaques in proximal and distal regions of BCA in apoE/LDLR-/-mice. In conclusion, OMSB staining represents a novel staining that could be routinely used for qualitative and quantitative microscopic assessments of formaldehyde-fixed and paraffin-embedded sections of arteries with atherosclerotic lesions.
Department of Histology, Jagiellonian University Medical College, Kopernika 7, 31-034, Kraków, Poland. mmgajda@cyf-kr.edu.pl.
Mariusz
http://orcid.org/0000-0002-3788-7528
M
Gajda
Jagiellonian Centre for Experimental Therapeutics (JCET), Jagiellonian University, Bobrzyńskiego 14, 30-348, Kraków, Poland.
Agnieszka
A
Jasztal
Jagiellonian Centre for Experimental Therapeutics (JCET), Jagiellonian University, Bobrzyńskiego 14, 30-348, Kraków, Poland.
Tomasz
T
Banasik
Department of Histology, Jagiellonian University Medical College, Kopernika 7, 31-034, Kraków, Poland.
Ewa
E
Jasek-Gajda
Jagiellonian Centre for Experimental Therapeutics (JCET), Jagiellonian University, Bobrzyńskiego 14, 30-348, Kraków, Poland.|Chair of Pharmacology, Jagiellonian University Medical College, 31-531, Kraków, Poland.
Stefan
S
Chlopicki
D000818:Animals; D001057:Apolipoproteins E / Q000172:deficiency*; D001391:Azo Compounds / Q000032:analysis* / Q000737:chemistry; D016122:Brachiocephalic Trunk / Q000473:pathology*; D051379:Mice; D018345:Mice, Knockout; D010078:Oxazines / Q000032:analysis* / Q000737:chemistry; D058226:Plaque, Atherosclerotic / Q000473:pathology*; D036301:Qualitative Research; D011973:Receptors, LDL / Q000172:deficiency*; D013194:Staining and Labeling
D001057:Apolipoproteins E; D001391:Azo Compounds; D010078:Oxazines; D011973:Receptors, LDL; C002313:PAcein; C042643:Martius scarlet blue trichrome
ApoE/LDLR−/− mice; Atherosclerosis; Elastic fibers; Martius scarlet blue; Orcein; Polychrome staining