Listeria monocytogenes is a Gram positive bacterial pathogen frequently used as a major model for the study of intracellular parasitism. Imaging late L. monocytogenes infection stages within the context of small-interfering RNA screens allows for the global study of cellular pathways required for bacterial infection of target host cells.

Genetics

Single-cell Gene Expression Using Multiplex RT-qPCR to Characterize Heterogeneity of Rare Lymphoid Populations

Thibaut Perchet¹, Sylvestre Chea¹, Milena Hasan², Ana Cumano¹, Rachel Golub¹

¹Unit for Lymphopoieseis, Immunology Department, INSERM U1223, University Paris Diderot, Sorbonne Paris Cité, Cellule Pasteur, Institut Pasteur, ²Center for Translational Science, Institut Pasteur, INSERM UMS20

This protocol describes how to assess the expression of a large array of genes at the clonal level. Single-cell RT-qPCR produces highly reliable results with a strong sensitivity for hundreds of samples and genes.

Biology

Analysis of Cell Suspensions Isolated from Solid Tissues by Spectral Flow Cytometry

Sandrine Schmutz¹, Mariana Valente^2,3,4,5, Ana Cumano², Sophie Novault¹

¹Flow Cytometry Core Facility, Center for Translational Research-Technical Core, Institut Pasteur, ²Unit for Lymphopoiesis, Immunology Department, INSERM U1223, University Paris Diderot, Sorbonne Paris Cité, Cellule Pasteur, Institut Pasteur, ³Stem-Cell Microenvironments in Repair/Regeneration Team, Instituto de Investigação e Inovação em Saúde (i3s), INEB - Instituto de Engenharia Biomédica, ⁴ICBAS - Instituto de Ciências Biomédicas Abel Salazar, Universidade do Porto, ⁵Stem Cells and Regenerative Medicine Team, UMRS 1166, ICAN - Institute of Cardiometabolism And Nutrition, UPMC - Université Pierre et Marie Curie - Paris 6, INSERM

This article describes spectral cytometry, a new approach in flow cytometry that uses the shapes of emission spectra to distinguish fluorochromes. An algorithm replaces compensations and can treat auto-fluorescence as an independent parameter. This new approach allows for the proper analysis of cells isolated from solid organs.

Biology

Isolation and Characterization of the Immune Cells from Micro-dissected Mouse Choroid Plexuses

Amaia Dominguez-Belloso¹, Sandrine Schmutz², Sophie Novault², Laetitia Travier^*1, Aleksandra Deczkowska^*1

¹Brain-Immune Communication Lab, Institut Pasteur, ²Cytometry platform, CB UTechS, Institut Pasteur

This study uses flow cytometry and two different gating strategies on isolated perfused mice brain choroid plexuses; this protocol identifies the main immune cell subsets that populate this brain structure.