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DUO82049 Sigma-Aldrich

Duolink® In Situ Wash Buffers, Fluorescence

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Properties

Related Categories Duolink PLA Accessories, Duolink PLA Technology, Molecular Biology, Protein Interaction, Proteomics More...
material   packing (powdered buffer pouches)
application(s)   proximity ligation assay: suitable
suitability   suitable for fluorescence
storage temp.   20-25°C

Description

Application

Duolink®proximity ligation assay(PLA®) allows for endogenous detection of protein interactions, post translational modifications, and protein expression levels at the single molecule level in fixed cells and tissue samples.

Use the Duolink® In Situ Fluorescence Protocol for this product. A set of short instructionsis also available.

Visit our Duolink® PLA Resource Center for information on how to run a Duolink® experiment, applications, troubleshooting, and more.

To perform a complete Duolink® PLA in situ experiment you will need two primary antibodies (PLA, IHC, ICC or IF validated) that recognize two target epitopes. Other necessary reagents include a pair of PLA probes from different species (one PLUS and one MINUS), detection reagents, wash buffers, and mounting medium. Note that the primary antibodies must come from the same species as the Duolink® PLA probes. Analysis is carried out using standard immunofluorescence assay equipment.

Specificity
Duolink® In Situ fluorescence applications use two wash buffers. Wash Buffer A is used after the PLA Probe incubation step and Wash Buffer B is used after incubation with the amplification reagents. See datasheet for more information.

Application Note
Two primary antibodies raised in different species 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®PLA 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). No minimum number of cells is required.

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Duolink® PLA 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).

Features and Benefits

• No overexpression or genetic manipulation required
• High specificity (fewer false positives)
• Single molecule sensitivity due to rolling circle amplification
• Relative quantification possible
• No special equipment needed
• Quicker and simpler than FRET
• Increased accuracy compared to co-IP
• Publication-ready results

Legal Information

Duolink is a registered trademark of Sigma-Aldrich Co. LLC

PLA is a registered trademark of Sigma-Aldrich Co. LLC


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Protocols & Articles

Articles

Duolink® PLA Applications

With Duolink® PLA, you can detect, quantify, and visualize protein-protein interactions, post-translational modifications, and low expression protein detection with an easy-to-execute immunodetection...
Keywords: Acylations, Amplification, Biological processes, Buffers, DNA replication, Electrophoresis, Flow cytometry, Gel electrophoresis, Gene expression, Glycosylations, High performance liquid chromatography, Immunohistochemistry, Mass spectrometry, Metabolism, Microscopy, Phosphorylations, Post translational modifications, Sulfations, Transcription, Transduction, Ubiquitinations

Duolink® References

Duolink proximity ligation assay (PLA) has been used in countless studies in a variety of fields. Browse the following tables to view highlights from the many applications of this technology.
Keywords: Apoptosis, Cancer, Cell biology, Coagulation, DNA replication, Degradations, Epigenetics, Flow cytometry, Gene expression, Growth factors, Immunology, Methylations, Neuroscience, Oxidations, PAGE, Peptide synthesis, Phosphorylations, Polymorphisms, Post translational modifications, Transcription

Protocols

Duolink® In Situ Short Instructions - Fluorescence

Note: Use open droplet reactions without a cover slip and perform all incubations in a humidity chamber. Use volumes corresponding to your reaction area, see the Reaction Volume Guide at sigma.com/du...
Keywords: Amplification, Confocal microscopy, Peptide synthesis, Phase transitions

Duolink® PLA Fluorescence Protocol

This protocol describes the use of Duolink® PLA reagents for the immunofluorescent detection, visualization, and quantification of individual proteins, protein modifications, and protein interactions...
Keywords: Amplification, Buffers, Confocal microscopy, Gene expression, Microscopy, Peptide synthesis, Titrations

Duolink® PLA Multicolor Detection Protocol

This protocol describes the use of Duolink® PLA Multicolor reagents for the simultaneous detection, visualization, and quantification of up to four proteins, protein modifications, and/or protein int...
Keywords: Amplification, Bacterial conjugations, Buffers, Confocal microscopy, Microscopy, Peptide synthesis, Phosphorylations

Duolink® PLA Probemaker Guide for Multicolor Detection

This guide describes the use of the Duolink® PLA Multicolor Probemaker kits for the conjugation of a unique pair of PLA oligonucleotides (Red Oligos A and B, Green Oligos C and D, Orange Oligos F and...
Keywords: Bacterial conjugations, Dialysis, Immunohistochemistry, Titrations

Duolink® PLA Product Selection Guide

Getting started with Duolink® PLA is simple. The protocols are very similar to that of IF, IHC, and flow cytometry, however Duolink® PLA offers distinct advantages over these and other traditional me...
Keywords: Amplification, Bacterial conjugations, Buffers, Flow cytometry, Fluorescent microscopy, Gene expression, Immunohistochemistry, Microscopy, Peptide synthesis

Duolink® PLA Troubleshooting Guide

The Duolink® PLA technology for protein detection offers a simple and straightforward protocol that has a set of defined steps. This section will delineate important reminders that will ensure your D...
Keywords: Addition reactions, Amplification, Bacterial conjugations, Buffers, Dehydration reaction, Filtration, Flow cytometry, Immunofluorescence, Immunohistochemistry, Peptide synthesis, Purification, Titrations

How to Optimize the Duolink® Proximity Ligation Assay

To successfully execute a Duolink® PLA experiment, there are some necessary considerations to properly prepare for, setup, and execute the assay protocol.
Keywords: Bacterial conjugations, Buffers, Detergents, Flow cytometry, Fluorescent microscopy, Gene expression, Growth factors, Immunofluorescence, Immunohistochemistry, In Situ hybridization, Microscopy, Nucleic acid hybridization, Phosphorylations, Post translational modifications, Sample preparations

Related Content

Duolink® PLA - Protein Detection Technology

Inside every cell, there is a breakthrough waiting to be uncovered. The chance to change the origin of a human disease or discover the tell-tale sign of an irregular protein-protein interaction requi...
Keywords: Amplification, Genetic, Immunofluorescence, Individual protein Immunoprecipitation, Peptide synthesis, Post translational modifications, Protein complex immunoprecipitation, Western blot

Duolink® PLA Custom Services

Duolink® Proximity Ligation Assay (PLA) is a powerful tool that allows in situ detection of endogenous proteins, protein interactions, and protein modifications with high specificity and sensitivity....
Keywords: Flow cytometry, Genetic, Peptide synthesis

Duolink® PLA Resource Center

Understanding protein function in the cell can create solutions to improve human health. For that reason we created the Duolink® PLA Resource Center, to provide insightful educational resources so yo...
Keywords: Flow cytometry, Peptide synthesis

Search Primary Antibodies for use with Duolink® PLA

The use of high quality, primary antibodies is critical to success for running a Duolink® Proximity Ligation Assay. We offer over 300 PLA-validated antibodies and antibody pair kits, in addition to t...
Keywords: Immunohistochemistry, PAGE, Peptide synthesis

Peer-Reviewed Papers
15

References

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