Digitally guided microdissection aids somatic mutation detection in difficult to dissect tumors

Abstract

Molecular genetic testing on formalin fixed, paraffin embedded (FFPE) tumors frequently requires dissection of tumor from tissue sections mounted on glass slides. In a process referred to as "manual macrodissection," the pathologist reviews an H&E stained slide at the light microscope and marks areas for dissection, and then the laboratory performs manual dissection from adjacent sections without the aid of a microscope, using the marked reference H&E slide as a guide. Manual macrodissection may be inadequate for tissue sections with low tumor content. We compared manual macrodissection to a new method, digitally guided microdissection, on a series of 32 FFPE pancreatic cancer samples. KRAS hotspot mutation profiling was performed using the Sequenom MassARRAY system (Agena Bioscience). Digitally guided microdissection was performed on multiple smaller areas of high tumor content selected from within the larger areas marked for manual macrodissection. The KRAS mutant allele fraction and estimated neoplastic cellularity were significantly higher in samples obtained by digitally guided microdissection (p < 0.01), and 7 of the 32 samples (22%) showed a detectable mutation only with digitally guided microdissection. DNA quality and yield per cubic millimeter of dissected tissue were similar for both dissection methods. These results indicate a significant improvement in tumor content achievable with digitally guided microdissection.