Description
Application
Based on proximity ligation assay (PLA), the Duolink® PLA Technology allows for endogenous detection of protein interactions, post-translational modifications, and protein expression levels at the single molecule level in fixed cells.
Duolink® Multicolor flow PLA Reagent Pack Kits used with Duolink® PLA Multicolor Probemaker Kits will enable Multiplexing of up to 4 protein events with sensitive detection of proteins, protein-protein interactions, and protein modifications within cell populations by flow cytometry. To perform a Duolink® flowPLA experiment, you will need fixed and permeabilized suspended cells, and at least two primary antibodies that specifically recognize your proteins of interest. Analysis is carried out using standard flow cytometry assay equipment. User must provide a fixed cell suspension and primary antibodies.
Application Note
Primary antibodies are needed. Test your primary antibodies (IgG-class, mono- or polyclonal) in a standard immunofluorescence (IF), immunohistochemistry (IHC) or immunocytochemistry (ICC) assay to determine the optimal fixation, blocking, and titer conditions. Duolink® in situ reagents are suitable for use on fixed cells, cytospin cells, cells grown on slide, formalin-fixed, paraffin embedded (FFPE), or tissue (fresh or frozen). Flow validated antibodies are recommended. It is also highly recommended that users purchase our Duolink(insert registration symbol) Compensation Beads (make title a hyperlink to beads products) to inform experimental design, instrument settings, and assist with compensation parameters.
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Biochem/physiol Actions
Fluorescence detection reagents are often used with FITC, Cy®3,Texas Red® or Cy®5 filter. The filter chosen is dependent on the color of the Multicolor Probemaker kit selected.
Duolink® flowPLA Multicolor Reagent Pack contains all the necessary reagents to perform the amplification and detection of bound custom PLA probes generated using Duolink® PLA Multicolor Probemaker Kits. Sufficient components are provided for approximately 40 tests at 100 μL reaction volume.
Features and Benefits
Visualize and detect multiple proteins events in one experiment
No overexpression or genetic manipulation required
High specificity (fewer false positives)
Detection of low abundant proteins due to rolling circle amplification
Relative quantification possible
No special equipment needed
Faster and more simple than FRET
Publication-ready results
Other Notes
This product is comprised of the following:
5x Multicolor PLA Ligation Buffer (DUO86002)
5x Amplification Buffer (DUO82050)
5x Multicolor FlowPLA Detection Buffer (DUO86003)
1x Ligase (DUO82025)
1x Polymerase (DUO82026)
1x Blocking Solution (DUO82007)
Duolink Probemaker PLA Probe Diluent (DUO82036)
See datasheet for more information.
Legal Information
Cy is a registered trademark of Cytiva
Duolink is a registered trademark of Merck KGaA, Darmstadt, Germany
Texas Red is a registered trademark of Life Technologies
