Duolink® PLA Control Kit - PPI
SIGMA/DUO92202 - Control Kit to reliably detect Protein-Protein Interactions (PPI) with Duolink® PLA
Synonym: in situ Proximity Ligation Assay Control Kit; Protein Protein Interaction Assay Control Kit
Product Type: Chemical
Schematic of a Duolink® PLA reaction. (A) Two primary antibodies recognize specific protein(s) of interest in the cell. The yellow and green globules represent two epitopes of a single protein or epitopes on two different proteins that are interacting. (B) Secondary antibodies coupled with oligonucleotides (PLA probes) bind to the primary antibodies. (C) When the PLA probes are in close proximity, connector oligos join the PLA probes and become ligated. (D) The resulting closed, circular DNA template becomes amplified by DNA polymerase. (E) Complementary detection oligos coupled to fluorochromes hybridize to repeating sequences in the amplicons. (F) PLA signals are detected by fluorescent microscopy as discrete spots and provide the intracellular localization of the protein or protein interaction.
Schematic of EGFR-HER2 dimerization. Ligands, such as epidermal growth factor (EGF) bind to the extracellular domain of EGFR, resulting in the activation and autophosphorylation. This, in turn, results in EGFR homodimerization (not depicted) or heterodimerization with other ErbB family members, such as HER2. Dimerization and subsequent phosphorylation lead to the activation of downstream signaling pathways.
Duolink®PLA Control Kit – PPI slide layout and sample placement. All wells contain EGF-stimulated, pre-fixed SK-OV3 cells. After permeabilization and blocking, wells 1 and 5 were incubated with both rabbit anti-EGFR and mouse anti-HER2 primary antibodies, wells 2 and 6 were incubated with anti-EGFR antibody alone, wells 3 and 7 were incubated with anti-HER2 antibody alone, and wells 4 and 8 were incubated with antibody diluent only.
| product line | Duolink® |
| shipped in | dry ice |
| storage temp. | −20°C |
| suitability | suitable for fluorescence |
| technique(s) | immunofluorescence: suitable |
| proximity ligation assay: suitable |
| Application: | Application note In the Duolink® PLA Control Kit, the cells and antibodies are provided, and the conditions have been optimized. When combined with the necessary Duolink® PLA reagents, you will have all you need to successfully perform the PLA experiment and obtain positive results. The Control kit contains Duolink® PLA Control Slides (Cat. No. DUO82202) - two 8 well glass slides with fixed SK-OV3 Human ovarian cancer cells. The cells were treated with EGF, to induce EGFR and HER2 interaction. The Kit also contains one vial of Anti-HER2 antibody produced in rabbit (HPA001383), and one vial of Monoclonal Anti-EGFR antibody produced in mouse (E3138) – the two primary antibodies you need to detect the two interacting protein Let us do the work for you, learn more about our Custom Service Program  to accelerate your Duolink® projectsView full Duolink® product list |
| 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® PLA Control Kit - PPI is designed to allows users to gain confidence with the Duolink® PLA technology. This Kit contains glass slides with fixed, treated cells and a specific pair of primary antibodies that will detect interacting proteins and reliably generate PLA signals when used with the complementary Duolink® PLA probes and detection reagents. Analysis is carried out using standard immunofluorescence assay equipment. Specificity This kit will enable the detection of Protein-Protein interactions. The SK-OV-3 cells contain HER2 and EGFR, which show increased interaction upon treatment with EGF. When combined with appropriate Duolink® PLA Reagents, the Control Kit will enable the successful detection of increased HER2 and EGFR protein interactions in the EGF treated cells. |
| Disclaimer: | RESEARCH USE ONLY. This product is regulated in France when intended to be used for scientific purposes, including for import and export activities (Article L 1211-1 paragraph 2 of the Public Health Code). The purchaser (i.e. enduser) is required to obtain an import authorization from the France Ministry of Research referred in the Article L1245-5-1 II. of Public Health Code. By ordering this product, you are confirming that you have obtained the proper import authorization. |
| Features and Benefits: | • Reliable PLA signal detecting a protein interaction • Gain confidence with the Duolink® PLA Technology • Add additional controls to your PLA experiments • Increased sensitivity due to rolling circle amplification for low abundant targets • No overexpression or genetic manipulation required • Relative quantification possible • Works with any fluorescence microscope and appropriate filters |
| General description: | Duolink® PLA Control Kits are intended for use with Duolink® PLA reagents to ensure proper function of reagents and Duolink® PLA staining technique. |
| Legal Information: | Duolink is a registered trademark of Merck KGaA, Darmstadt, Germany |
| Legal Information: | PLA is a registered trademark of Merck KGaA, Darmstadt, Germany |
| Other Notes: | Other Duolink® reagents needed (30RXN size): Mouse/Rabbit Duolink® PLA Starter Kit: Red Starter Kit (DUO92101 ) OR Orange Starter Kit (DUO92102 )OR PLA Probe pairs (PLUS and MINUS): Anti-Mouse PLUS (DUO92001 ) and Anti-Rabbit MINUS (DUO92005 ) or Anti-Mouse MINUS (DUO92004 ) and Anti-Rabbit PLUS (DUO92002 ); AND Detection reagents of choice: Orange (DUO92007 ), Red (DUO92008 ), FarRed (DUO92013 ), or Green (DUO92014 ); AND Wash Buffers (DUO82049 ) AND Mounting Medium with DAPI (DUO82040 ). |
| Other Notes: | This product is comprised of the following: • Duolink® PLA Control Slides (DUO82202) - two 8-well chamber slides with EGF treated, pre-fixed SK-OV3 cells. • One vial (15μL) of Rabbit anti-ErbB2/HER2 antibody (HPA001383 ) • One vial (25μL) of Mouse anti-EGFR antibody (E3138 )See datasheet for more information. |
| RIDADR | NONH for all modes of transport |
| Storage Temp. | −20°C |
| UNSPSC | 41105331 |