We use a counterstain, called Acid Blue 129, that our team developed several years ago (Zehntner et al., J. Histochem. Cytochem., 56: 873-880, 2008; doi:10.1369/jhc.2008.950345). This stain has a homogenous light blue background that provides excellent contrast to the red-brown chromogen and is well-suited for image segmentation and quantitative analysis.