Abstract 2590: Automating 6-plex chromogenic multiplexing with BOND RX

Abstract

Abstract Multiplex staining using a sequential approach provides a means of visualizing multiple targets in a single biopsy section, further evaluation paired with digital imaging provides a deeper analysis into the spatial relationship at a cellular level. Leica Biosystems enables researchers to complete proof of concept for sequential multiplex staining methods, on board the BOND RX system. Thus, paired with a compatible reagent workflow and a digital slide imaging offering (Aperio GT 450/Aperio AT2) enhances the possibilities for analysis. As with any assay there is a requirement for user optimization, in particular, marker/chromogen layer selection and the pertinent use of an elution step. Such elution step is specifically important within a sequential stain utilizing the same detection reagents on subsequent layers. The results from the multiple staining rounds show a successful 6-plex fully automated stain, possible without the removal and reapplication of the slide onto the BOND system between staining layers. The marker combinations chosen demonstrate compatibility of different host species (Rabbit & Mouse), cellular compartments and isotypes which are provided by Leica Biosystems as BOND Ready to Use reagents. Overall, chromogenic multiplexing provides a stained image which enables color separation, due to the assignment of each chromogen to a marker with an individual cell type and cellular compartment. Finally, the process outlined avoids the risk of signal quenching commonly associated with fluorescent offerings and negates the requirement for specialized equipment due to the enablement of brightfield primary analysis. Citation Format: Dean Talia, Stacey Lindsay, Sarah Ketting, Rebecca Williamson, Damian Cockfield. Automating 6-plex chromogenic multiplexing with BOND RX [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 2590.