Staining methods for Calcium—a Comparison Based on Reactions Shown by Cardiac Calcinosis in DBA Mice

Spontaneous calcification, which occurs frequently in hearts of mice of the DBA strain, has been used to compare effectiveness of calcium stains. Three fixatives (1) 80% ethanol, (2) 10% formalin and 3.5% acetic acid in 95% ethanol (modified Lavdowsky's) and (3) a 1:1 mixture of concentrated formalin and absolute ethanol were tested for their subsequent effects on staining. Stains for Ca included purpurin, von Kossa's, phthalocyanin, and glyoxal bis(2-hydroxyanil). These, with appropriate counterstaining, were compared among each other, with hematoxylin-eosin (H-E), and with the periodic acid-Schiff (PAS) reaction. Fixatives (1) and (3) did not affect results appreciably, but the von Kossa technic was unsatisfactory after (2). All stain demonstrated sites of Ca deposits, but purpurin and von Kossa's gave greatest sharpness of definition and were the simplest to perform. H-E and PAS with Alcian blue showed the same Ca sites in addition to details of the surrounding tissue.