Exosome Isolation and Analysis Services Offered By ResearchDx
- qPCR Expression Analysis
- miRNA NGS expression analysis
- Total RNA NGS expression analysis
- Total DNA analysis via NGS
- ELISA-based protein analysis
- Fluorescence-activated Cell Sorting (FACS)
- Flow Cytometry
High Throughput Exosome Extraction
- DNA
- RNA
- MiRNA
- Protein
How Does EXO-NET® work?
EXO-NET is a fast and efficient exosome isolation system suitable for almost any biofluid or application. The EXO-NET workflow can be applied to high throughput screening of EVs and exosome-based biomarker discovery for diagnostics and therapeutic targets.
What are Exosomes
Exosomes are membrane-bound extracellular vesicles (EVs) produced in most eukaryotic cells’ endosomal compartments. In multicellular organisms, exosomes and other EVs are found in biological fluids, including saliva, blood, urine, and cerebrospinal fluid. EVs have specialized functions in physiological processes, from coagulation and waste management to intercellular communication.
Exosomes are smaller than most other EVs, from about 30 to 150 nanometres (nm) in diameter, around the same size as many lipoproteins but much smaller than cells.
EVs in circulation carry genetic material and proteins from their cell of origin, proteo-transcriptomic signatures that act as biomarkers. In the case of cancer cells, exosomes may show differences in size, shape, morphology, and canonical markers from their donor cells. They may encapsulate relevant information that can be used for disease detection. Consequently, there is a growing interest in clinical applications of EVs as biomarkers and therapies.
EXO-NET Performance
Comparison of mRNA and microRNA yield and recovery from EXO-NET isolated EVs with 4 other EV isolation kits.
Method: EVs were isolated from pooled normal human plasma by EXO-NET and other 4 commercial kits according to their manufacturers’ instructions. Isolated EVs were lysed and total RNA was extracted for qPCR analysis to measure both microRNAs (miR-16, let-7a and miR-21) and mRNAs (GAPDH, OAZ1, RPLPO and SERF2).
Result: EXO-NET delivers equivalent or higher recovery of plasma EV RNA (mRNAs and microRNAs) compared to the other 4 commercial EV isolation kits as indicated by a lower CT value
Evaluation of known EV protein and RNA markers in EXO-NET isolated EVs from human plasma.
Method: Pooled normal human plasma (500µl) was incubated with 30 µl of EXO-NET for 15 min. EXO-NET-captured EVs were magnetically isolated and lysed for downstream analysis. Total EV protein and RNA were extracted and analysed by Western blot and qPCR.
Result: Western blot analysis showed EXO-NET-isolated EVs from normal human plasma were positive for CD63, CD81, CD9, TSG101 and Flotillin-1, and negative for calnexin (an EV negative marker). qPCR analysis demonstrated that EXO-NET isolated EVs had high yield of mRNA (GAPDH) and microRNA (miR-191).
Comparison of known EV proteins and RNA markers in EXO-NET isolated EVs from human saliva.
Method: Pooled normal human saliva (400µl) was diluted with 400µl of PBS and then incubated with 30 µl of EXO-NET for 15 min. EXO-NET captured EVs were magnetically isolated and lysed for downstream analysis. Total protein and RNA were extracted from captured EVs and analyzed by Western blot and qPCR.
Result: Western blot analysis confirmed the presence of CD63, CD81 and CD9. Calnexin (EVs negative marker) was not detectable. qPCR analysis confirmed that EXO-NET isolated EVs had a high yield of mRNA (GAPDH) and microRNA (U6).
Comparison of mass spectrometry proteomic analysis of EXO-NET isolated EVs with another bead-based EV isolation kit.
Method: EVs were isolated from pooled normal human plasma by EXO-NET and Kit A according to manufacturers’ instructions. Isolated EVs were lysed and extracted total protein were processed for proteomic analysis by tandem mass spectrometry.
Result: Mass spectrometry analysis showed that albumin peptide intensity in Kit A was 2-fold higher than that observed for EXO-NET isolated EVs. EXO-NET reduces contamination of EV preparations by serum proteins that may confound downstream analysis. The EV peptides intensity in Kit A was 6 to 10-fold less than that observed for EXO-NET-isolated EVs.
ResearchDx is now offering EXO-NET® Exosome Isolation and Analysis services through a licensing agreement with Inoviq