Introduction

Obtaining a definitive diagnosis of canine histiocytic sarcoma (HS) can be challenging. Two proteins, IBA-1 and CD204, are immunohistochemical markers helpful in distinguishing HS from other tumor types with similar morphological features. This study was performed to validate the use of a novel RNA In Situ Hybridization (ISH) technology (RNA-ISH) allowing single-molecule RNA visualization in Formalin‐Fixed Paraffin‐Embedded (FFPE) tissues and cell pellets as a diagnostic tool for the diagnosis of canine HS.

Methods

RT-PCR, genetic sequencing, Western Blot analysis and immunohistochemistry for IBA-1 and CD204 were performed to correlate gene expression and protein expression in the DH82 canine HS cell line. Once validated in the DH82 cell line, RNA-ISH for IBA-1 and CD204 was performed in clinical FFPE HS samples to demonstrate mRNA expression of these markers. FFPE archived samples of various round cell tumors (lymphomas, mast cell tumors, and plasma cell tumors) and melanomas were used as negative control for RNA-ISH against IBA-1 and CD204.

Results

RNA-ISH showed moderate to strong expression of RNA for IBA-1 and CD204 genes in DH82 cell line and FFPE canine HS samples. RNA-ISH showed little to no expression of the target genes in control samples. The minimal expression of CD204 and IBA-1 seen in these tumors was assessed to be secondary to tumoral macrophagic infiltrates.

Conclusion

RNA-ISH for IBA-1 and CD204 was highly specific and sensitive. It may be a valuable diagnostic technique in the diagnosis of canine HS.

Funding Information

The Finn Project provided financial support for this study, as well as the Florida Veterinary Scholars Program and Boehringer Ingelheim Veterinary Research Scholars Program.