Abstract 3868: High throughput tissue phenotyping and imaging of the tumor immune microenvironment using novel FlexVUE™ multiplexed immunofluorescence assays

Abstract

Abstract Immunotherapy has transformed the treatment of metastatic and recurrent solid tumors. Advances in technology in the past few years have created unprecedented opportunities to identify biomarkers of disease processes, especially by using multi-omics technologies and datasets to derive valid and useful signatures of disease. Advances in multiplexed immunofluorescence (mIF) staining and imaging technologies that allow for accurate phenotyping of individual cells while also preserving architectural features of the sample are critical to our understanding of the interplay of tumor and immune cells in the tumor immune microenvironment (TiME).Herein we describe the utility of our new flexible mIF assays (FlexVUE™ panels) to provide the necessary relevant distribution of infiltrating immune cells in tumors coupled to the use of our new UltiStacker™ software. This hardware and software package allows a detailed spatial characterization of specific cell phenotypes defined by co- or lack of expression that may help in predicting clinical responses and mechanisms of resistance to therapy. The FlexVUE™ mIF assay was used to conduct 8-plex immunophenotyping of FFPE tumor sections. Slides were stained with a cocktail of eight primary antibodies using Leica Biosystems BOND RX autostainer and imaged on the Zeiss Axio Scan.Z1. Following the first round of imaging, the slides were decoverslipped and the signal was removed. New targets were probed on the same tissue section in a reagent incubation step on the BOND RX, termed ‘exchange.’ Slides were re-imaged on the Zeiss Axio Scan Z1 using the same dye channels. The image pairs were automatically aligned using UltiStacker™ software and exported for downstream analysis with Indica Labs HALOv3.1 software. Our approach demonstrates a streamlined off the shelf workflow requiring no assay development time that supports whole slide imaging of a flexible high plex panel. In addition, our assays enable traditional same slide H&E fusion on a single tissue slide for comprehensive tissue immunophenotyping analysis of proliferating cells, tumor cells, tumor-infiltrating lymphocytes, macrophages or tertiary lymphoid structures. Citation Format: Yvette Cajigas, Peter Hamer, Alina Ainbinder, Angela Vasaturo, Douglas Wood, Kirsteen H. Maclean, Gourab Chatterjee. High throughput tissue phenotyping and imaging of the tumor immune microenvironment using novel FlexVUE™ multiplexed immunofluorescence assays [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3868.