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Detachment Procedure of Bacteria from Atmospheric Particles for Flow-cytometry Counting

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Flow Cytometry Assay for Recycling of LFA-1 in T-lymphocytes

Authors: Katarzyna Potrzebowska, Janne Lehtonen, Malin Samuelsson and Lena Svensson, date: 12/05/2018, view: 1031, Q&A: 0

[Abstract] To enable cells to move forward, cell surface integrins are internalized into an endosomal compartment and subsequently intracellularly transported to be re-exposed at a new site on the cell membrane. Leukocytes are the fastest migrating cell type in the human body, which express the leukocyte-specific integrin LFA-1. Here, we describe a flow ...

Identification and Quantitation of Leukocyte Populations in Human Kidney Tissue by Multi-parameter Flow Cytometry

Authors: Katrina Kildey, Becker M.P. Law, Kimberly A. Muczynski, Ray Wilkinson, Helen Healy and Andrew J. Kassianos, date: 08/20/2018, view: 1616, Q&A: 0

[Abstract] Inflammatory immune cells play direct pathological roles in cases of acute kidney injury (AKI) and chronic kidney disease (CKD). However, the identification and characterization of distinct populations of leukocytes in human kidney biopsies have been confounded by the limitations of immunohistochemical (IHC)-based techniques used to detect them. ...

Visualization of RNA at the Single Cell Level by Fluorescent in situ Hybridization Coupled to Flow Cytometry

Authors: Alice Coillard and Elodie Segura, date: 06/20/2018, view: 2240, Q&A: 0

[Abstract] The protocol described here has been developed to detect RNA at the single cell level. Fluorescent probes hybridize to target RNAs and are detected by flow cytometry after multiple amplification steps. Different types of RNA can be detected such as mRNA, long noncoding RNA, viral RNA or telomere RNA and up to 4 different target probes can be used ...

Analysing Temporal Dynamics of T Cell Division in vivo Using Ki67 and BrdU Co-labelling by Flow Cytometry

Authors: Thea Hogan, Andrew Yates and Benedict Seddon, date: 12/20/2017, view: 4465, Q&A: 0

[Abstract] This protocol was developed to increase the richness of information available from in vivo T cell proliferation studies. DNA labelling techniques such as BrdU incorporation allow precise control of label administration and withdrawal, so that the division history of a population can be tracked in detail over long timeframes (days-weeks). ...

MHC Class II Tetramer Labeling of Human Primary CD4⁺ T Cells from HIV Infected Patients

Authors: Moran Galperin, Daniela Benati, Mathieu Claireaux, Madhura Mukhopadhyay and Lisa A. Chakrabarti, date: 03/20/2017, view: 4825, Q&A: 0

[Abstract] Major Histocompatibility Complex (MHC) tetramers have been used for two decades to detect, isolate and characterize T cells specific for various pathogens and tumor antigens. In the context of Human Immunodeficiency Virus (HIV) infection, antigen-specific CD8⁺ T cells have been extensively studied ex vivo, as they can be ...

Isolation of Highly Pure Primary Mouse Alveolar Epithelial Type II Cells by Flow Cytometric Cell Sorting

Authors: Meenal Sinha and Clifford A. Lowell, date: 11/20/2016, view: 11841, Q&A: 0

[Abstract] In this protocol, we describe the method for isolating highly pure primary alveolar epithelial type II (ATII) cells from lungs of naïve mice. The method combines negative selection for a variety of lineage markers along with positive selection for EpCAM, a pan-epithelial cell marker. This method yields 2-3 x 10⁶ ATII cells per mouse ...

Flow Cytometry of Lung and Bronchoalveolar Lavage Fluid Cells from Mice Challenged with Fluorescent Aspergillus Reporter (FLARE) Conidia

Authors: Anupam Jhingran, Shinji Kasahara and Tobias M Hohl, date: 09/20/2016, view: 7983, Q&A: 0

[Abstract] Aspergillus fumigatus is a ubiquitous fungal pathogen that forms airborne conidia. The process of restricting conidial germination into hyphae by lung leukocytes is critical in determining infectious outcomes. Tracking the outcome of conidia-host cell encounters in vivo is technically challenging and an obstacle to understanding ...

In vivo OVA-specific Cytotoxic CD8⁺ T Cell Killing Assay

Authors: Nada Chaoul, Catherine Fayolle and Claude Leclerc, date: 06/20/2016, view: 8718, Q&A: 0

[Abstract] Cytotoxic CD8⁺ T cells are responsible for the lysis of cells expressing peptides associated with MHC class I molecules and derived from infection with a pathogen or from mutated antigens. In order to quantify in vivo this antigen-specific CD8⁺ T cell killing activity, we use the in vivo killing assay (IVK). ...

In vitro Assessment of Immunological Synapse Formation by Flow Cytometry

Authors: Bo-Ra Na and Chang-Duk Jun, date: 03/20/2016, view: 9064, Q&A: 2

[Abstract] In adaptive immune system, formation of immunological synapse between T cells and antigen presenting cells (dendritic cells, B cells, and macrophages) or target cells (tumor cells and viral-infected cells) is critical for the execution of T cell immune responses via cytokine secretion or direct killing activity. Here, we describe the practical ...

Ex vivo Human Natural Killer (NK) Cell Stimulation and Intracellular IFNγ and CD107a Cytokine Staining

Authors: Vanessa A. York and Jeffrey M. Milush, date: 06/20/2015, view: 9404, Q&A: 1

[Abstract] Natural killer (NK) cells comprise 5–20% of peripheral blood mononuclear cells (PBMC) in humans. In addition to their fundamental roles in the defense against viral infections and tumor surveillance, NK cells help shape adaptive immune responses through their production of cytokines. NK cells are traditionally identified as CD3^neg, CD14 ...

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