Sample Preparation Optimization
The first step to obtaining high quality flow data is sample preparation. The critical step of preparing a single cell suspension will help to avoid unwanted instrument clogs and low quality data. Use this guide for tips on how to optimize your sample preparation method and start getting better results.
Keywords: flow cytometry optimization, sample preparation, FACS experiment sample prep
Permeabilization and Fixation Method Optimization
Various permeabilization and fixation methods are available to allow access of antibodies to intracellular proteins. Fixing cells will retain the target protein in its original cellular location, and will usually ensure better stability of soluble antigens and antigens with a short half-life. Use this guide to help decide which method is optimal for your experiment.
Keywords: FACS, flow cytometry, optimization, permeabilization, fixation method, fixative
Experimental Controls Optimization
Flow experiments can become very complex with multiple parameters and mixed cell populations, and sometimes there can be numerous experimental controls involved. Use this guide to help clarify and choose appropriate controls for your FACS experiment.
Keywords: flow cytometry, FACS experimental controls, negative and positive control, isotype, FMO controls
Fluorescent Conjugates (Fluorophore) Optimization
There are many fluorescent molecules, also known as fluorochromes, fluorophores, or fluorescent dyes, with a potential application in flow cytometry. By conjugating (pre-attaching) them to primary antibodies, we can create conjugated antibodies that allow for flow cytometry analysis. Use this guide to learn how to choose the right fluorescent labels for your FACS experiment.
Keywords: Fluorochrome selection, fluorophores, fluorescent dyes, labels, tags, molecules, fluorescent stain
Cell Stimulation Optimization (for Intracellular Cytokine Staining)
To optimize your flow experiment, additional in vitro cell stimulation may be required to trigger increased production of intracellular cytokines. Some common reagents for this purpose include PMA, Ca++ or peptide epitopes and protein transport inhibitor, Brefeldin A, and more. Use this guide to learn the optimal method for stimulating your cell samples before flow cytometric analysis.
Keywords: optimize in vitro cell stimulation, increase intracellular cytokine response
Antibody Immuno-Staining Optimization
The antibody staining procedure includes many steps, such as antibody titration, washing, blocking, and more. The optimal antibody concentration, which gives the best staining with minimum background, must be determined experimentally. Use this guide for tips on how to optimize the steps of your antibody staining protocol.
Keywords: FACS experiment optimization, antibody staining, titration, fluorescent staining