Comparison of collagen I and collagen III immunohistochemistry with Herovici staining in various rabbit organs.

Collagen I and III distribution is not only crucial to assess the status of healing wounds, but also to characterise healthy connective tissue and pathological extracellular matrix composition. In this technical note, we have therefore compared the dual-coloured Herovici staining, indicating pink collagen I and blue collagen III in serial sections with immunohistochemistry (IHC) labellings for collagen I and III, respectively. Furthermore, we used, chromogenic DAB for IHC labelling. Seven different organs of a healthy New Zealand white rabbit were collected for this purpose, including kidney, liver, tonsil, tongue, duodenum, heart, and brain, respectively. A dual-coloured staining like Herovici turned out to be as good as two single-colour labellings utilising IHC. In some cases, co-localisation and extent of collagen I and III expression could be qualitatively visualised better using Herovici, with gradients of blue-violet-pink, than by mere comparison of labelling intensities side by side in two different sections, although taken at the same place as serial sections. Nevertheless, a quantitative analysis of the Collagen I-to-III ratio revealed no significant differences between these two approaches to assess the extracellular matrix composition. From these comparisons, we conclude that a Herovici staining is recommended as a valuable alternative staining to collagen I and III IHC; and it may act as a fast and cheap preliminary staining method. These findings encourage researchers focusing on ECM composition of the experimental rabbit tissue to use Herovici staining to determine the ratio of the extracellular collagen I and III expression.