The Immune Analysis Facility (IAF), part of the Center for Immunotherapy at Roswell Park Comprehensive Cancer Center, is a shared resource responsible for serial monitoring of immunologic functions in patients with cancer, those who are treated with biologic therapies, and those who participate in clinical trials or research protocols. The IAF makes available to its users a broad range of state-of-the-art immunologic assays, performed under a rigorous quality control program. In addition, as advances in immunobiology occur and new assays are requested by the users, the IAF performs pre-clinical evaluations of the assays and, when they become reliable and standardized, adds them to the available assay list. The IAF offers consultation regarding optimal immunologic assessment and assay development for innovative approaches to evaluate immune responses. They provide expert advice regarding the types of assays for immune monitoring and data interpretation tailored to fit the endpoints of each specific clinical trial.
The IAF maintains extensive quality control (QC) and quality assurance (QA) programs to ensure the validity of test results.
Seahorse Extracellular Flux Analyzer
The Seahorse Extracellular Flux Analyzer is housed at the Roswell Park Center for Immunotherapy. This technology simultaneously measures oxygen consumption rate (OCR) and extra cellular acidification rate (ECAR), indicators of mitochondrial respiration and glycolysis respectively, in a microplate, in real-time. This instrument allows us to study substrate utilization, mitochondrial function, energy expenditure, bioenergetics health of cells, and the shift between the two pathways, oxidative phosphorylation (OXPHOS) and glycolysis. The OCR/ECAR ratio can determine dominance of glycolysis or OXPHOS as the cells differentiate. Real-time OCR is determined during sequential treatments with oligomycin, FCCP, and antimycin-A/rotenone supplied by Mito-Stress test kit, and (ECAR) is determined by glucose, oligomycin and 2-DG treatments which are the components of Glyco-Stress test kit. By using the Mito Stress test kit, we can determine maximal respiration and spare respiratory capacity, an indicator of mitochondrial function. By using Glyco Stress test kit, we can examine the ability of cells to undergo rapid glycolytic switch upon stimulation, a hallmark of functional T cell effectors. Investigators can use this technology to assess metabolic changes of cancer and immune cells to various clinically relevant conditions, including drug exposure and hypoxia.
A Helios/CyTOF is a time-of-flight mass cytometry which is capable of acquiring single cell suspension labeled with transition metal isotopes-conjugated monoclonal antibodies. The main advantages of mass cytometry over conventional fluorescence-based flow cytometry are minimal spectral overlap of each parameter, thereby the CyTOF can theoretically detect up to 100 isotopes without compensation. In addition, since the metal isotopes used for conjugation are rare in biological samples and environment, it can detect the specific signal of each protein on individual cell in the absence of autofluorescence. Overall this system greatly enhances the ability of researchers to simultaneously analyze individuals cells with >35 parameters for phenotype and function in a single tube.