Sandrine_Schmutz

Sandrine Schmutz

Flow Cytometry Core Facility - Pasteur Institut

Paris, France

14:25 - 14:55

Day 2 – February 6, 2025

From small particles to high dimensional analysis: various applications on the Bigfoot spectral sorter

Spectral flow cytometry is a powerful technique used to analyze multiple parameters of individual cells. In our core facility, we have evaluated the impact of spectral cytometry for various sorting applications that will be shown here.
Detecting and sorting Extracellular Vesicles (EVs) and Viruses (range of diameter from 40 to 130 nm) using spectral flow cytometry involves specific considerations due to their small size and heterogeneous nature. To overcome these challenges, we employed the BigFoot technology (ThermoFisher) with optimized parameters for the detection of sub-cellular particles, sample labeling, laser power, voltage settings combined with an adapted fluidic pathway. We have compared results obtained with the nanoFCM (nanofcm) and the ID7000 (Sony) technologies by using Fluorescent Tagging Beads of different size and composition.
On another side, spectral cytometry has proven its power in high dimensional single cell analysis, offering unprecedented capabilities for panel design and data resolution. Using default settings and automated unmixing algorithms, we assessed the performance and reproducibility of the BigFoot cell sorter on a 20-color immunophenotyping panel.
Our results highlight the significant advantages of spectral cytometry, including reduced complexity in panel set up and streamlined workflow due to the automation of spectral unmixing. Comparative analysis revealed consistent signal resolution and separation, validating the robustness of spectral technologies for analysis and sorting applications in both sub-cellular and multiparametric analysis.