Detection of Extracellular Vesicles Using Proximity Ligation Assay with Flow Cytometry Readout—ExoPLA
| Autor: | Linda Arngården, Tonge Ebai, Masood Kamali-Moghaddam, Liza Löf, Ulf Landegren, Ola Söderberg |
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| Rok vydání: | 2017 |
| Předmět: |
0301 basic medicine
In situ Histology Ligase Chain Reaction Proximity ligation assay Biology Biochemistry Extracellular vesicles Antibodies Flow cytometry Extracellular Vesicles 03 medical and health sciences medicine Animals Humans DNA Primers medicine.diagnostic_test General Medicine Flow Cytometry Fluorescence Molecular biology Microvesicles Medical Laboratory Technology 030104 developmental biology Rolling circle replication Biophysics |
| Zdroj: | Current Protocols in Cytometry |
| ISSN: | 1934-9300 1934-9297 |
| Popis: | Extracellular vesicles (EVs) are continuously released by most cells, and they carry surface markers of their cells of origin. Found in all body fluids, EVs function as conveyers of cellular information, and evidence implicates them as markers of disease. These characteristics make EVs attractive diagnostic targets. However, detection and characterization of EVs is challenging due to their small size. We've established a method, called ExoPLA, that allows individual EVs to be detected and characterized at high specificity and sensitivity. Based on the in situ proximity ligation assay (in situ PLA), proximal oligonucleotide-conjugated antibodies bound to their targets on the surfaces of the EVs allow formation of circular products that can be fluorescently labeled by rolling circle amplification. The intense fluorescent signals produced in this assay allow detection and enumeration of individual EVs by flow cytometry. We describe the procedures for ExoPLA, along with expected results and troubleshooting. © 2017 by John Wiley & Sons, Inc. Keywords: ExoPLA; exosomes; extracellular vesicles; flow cytometry; proximity ligation assay |
| Databáze: | OpenAIRE |
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