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Monitoring the Rab27 associated exosome pathway using nanoparticle tracking analysis (CAT#: STEM-MB-0708-WXH)

Introduction

Exosomes are secreted by many cell types and display multiple biological functions. The ability to both rapidly detect and quantify exosomes in biological samples would assist in the screening of agents that interfere with their release, and which may therefore be of clinical relevance. Nanoparticle tracking analysis, which detects the size and concentration of exosomes, was used to monitor the inhibition of exosome secretion from MDA-MB-231 breast cancer cells expressing inhibitory RNA targeted for Rab27a, a known component of the exosome pathway. Inhibition of both Rab27a and Rab27b was observed, resulting in alterations to intracellular CD63+ compartments and the release of fewer exosomes into the culture medium, as determined by nanoparticle tracking analysis and confirmed by immunoblotting and protein quantification.




Principle

Nanoparticle tracking analysis (NTA) is a method for visualizing and analyzing particles in liquids that relates the rate of Brownian motion to particle size. The rate of movement is related only to the viscosity and temperature of the liquid; it is not influenced by particle density or refractive index. NTA allows the determination of a size distribution profile of small particles with a diameter of approximately 10-1000 nanometers (nm) in liquid suspension.

Applications

NTA has been used by commercial, academic, and government laboratories working with nanoparticle toxicology, drug delivery, exosomes, microvesicles, bacterial membrane vesicles, and other small biological particles, virology and vaccine production, ecotoxicology, protein aggregation, orthopedic implants, inks and pigments, and nanobubbles.

Procedure

1. Sample preparation
2. Nanoparticle Tracking Analysis
3. NTA Data Processing, Display, and Interpretation

Materials

• Nanoparticle Tracking Analysis (NTA) instrument
• NanoSight instrument
• Nanoparticle Analyzer
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