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Flow Cytometry Protocols

Sample Preparation

 

 

Staining cells with a Lyse/No-Wash protocol


Direct Immunofluorescence Staining of Whole Blood Using a Lyse/No-Wash Procedure

Find procedures for whole blood specimen collection and preparation, including step-by-step instructions for lysing and staining. Use this method to detect cells bearing specific membrane antigens.

 

BD™ Phosflow Protocols for Mouse Splenocytes or Thymocytes

Explore four protocols using different methods (detergent method, mild or harsh alcohol method or harsh detergent method) for the activation of cell signaling molecules in mouse splenocytes or thymocytes.

 

 

Indirect Staining Mononuclear Cells


Indirect Immunofluorescence Staining of Mononuclear Cells

The indirect method is used to enhance the fluorescence signal and also to facilitate multicolor staining of human cells when direct conjugated reagents are not available. Find the procedure for specimen collection, preparation and staining of mononuclear cells.   
  

BD™ Phosflow Protocols for Human PBMCs

Explore four protocols using different methods (detergent method, mild or harsh alcohol method or harsh detergent method) for the activation of cell signaling molecules in  human peripheral blood mononuclear cells.

 

BD™ Phosflow Protocols for Human Whole Blood Samples

Explore four protocols using different methods (detergent method, mild or harsh alcohol method or harsh detergent method) for the activation of cell signaling molecules in  human whole blood samples.

 

 

Reducing non-specific staining with Fc Block


Uses of Fc Block

Learn how antibody binding to Fc receptor can cause background staining and how to use the BD Fc Block™ Reagents in immunophenotyping of mouse or rat leucocytes..   

  

Protocol for Direct Immunofluorescence Staining of Mononuclear Cells

Find the procedure for specimen collection, preparation and direct immunofluorescence staining of mononuclear cells..

 

Cell Surface Staining of Human PBMCs and Suspension Cell LinesD™ Phosflow Protocols for Human Whole Blood Samples

Find a step-by-step procedure for cell surface staining of human peripheral blood mononuclear cells and cell lines.

 

Phosflow Protocol for Adherent Cells

Find the optimal protocol for detecting intracellular signaling molecules from adherent cells using BD Phosflow™ Reagents.

 

Intracellular Flow and Phosflow FAQ

Find answers to most commonly asked questions on intracellular flow cytometry and use of BD Phosflow™ Reagents..

 

TCR Stimulation and 96 Well BD Phosflow Protocol

Find the complete procedure for TCR stimulation of peripheral whole blood cells using BD Phosflow buffers.

 

 

Controls


Isotype Control Antibody Selection Guide

The selection of an appropriate isotype control is essential for every flow cytometry experiment. Find procedures for determining the isotype of your primary antibody, for searching for your isotype control and for using your isotype control.

 

 

Troubleshooting
 

The Escapee Phenomenon

Escapees are aggregates of cells (including positively labeled cells) that are excluded from the flow cytometric analysis gate, thus altering the results of the analysis. See recommendations to decrease the number of escapees.

 

 

Intracellular Cytokine Staining Protocol


Immunofluorescent Staining of Intracellular Cytokines for Flow Cytometric Analysis

Get information on stimulation of cells, appropriate cultures for generating human, mouse and rat cytokine producing cells and describes a protocol for multicolor staining of intracellular cytokines and cell surface antigens. It also offers an alternative protocol for the activation and intracellular staining of whole blood.


Cytokine Stimulation and 96 Well BD Phosflow

Explore the complete procedure for cytokine stimulation of peripheral whole blood cells using BD Phosflow buffers.