Overview

A better way to isolate exosomes

“We therefore pursued the ExoQuick^® method for further study, as these samples required much less sample input, a key benefit when working with clinical samples and mouse models¹.” Need exosomes? SBI’s ExoQuick-TC enables high-throughput, quantitative isolation of exosomes from low volumes (as little as 1 ml) of tissue culture media and certain biofluids (saliva, urine, follicular fluid, and breast milk). Note, to isolate exosomes from serum, plasma, or ascites fluid, use the original ExoQuick formulation (EXOQ5A-1 or EXOQ20A-1)). Compatible with a wide variety of downstream applications, ExoQuick-TC is an effective and proven alternative to ultracentrifugation^1-3.

ExoQuick-TC’s fast, ultracentrifugation-free method:

• Saves time and labor
• Is easily scalable
• Conserves precious sample
• Delivers high yields of functional, high quality exosomes
• Can be used to isolate exosomes for a wide range of downstream applications, including
  • Biomarker studies
  • Exosomal miRNA profiling
  • Exosomal proteomics
  • Exosomal lipidomics/metabolomics
  • Functional studies, such as in cell-to-cell signaling
  • Basic biology, such as role in tumorigenesis

ExoQuick-TC is a proprietary polymer that gently precipitates exosomes. First, pre-clear your samples of cells and cellular debris, and then simply add the appropriate amount of ExoQuick-TC to your cleared biofluid, refrigerate, and centrifuge (see the product manual for protocol details). Your exosomes will be in the pellet, ready for resuspension in an appropriate solution.

In electron microscopy studies, exosomes isolated with ExoQuick-TC appear similar to exosomes isolated using ultracentrifugation^1-2, and these exosomes are also active in numerous functional assays^1-3.

Exosomes isolated with ExoQuick-TC can be used for all types of protein profiling and protein characterization studies, such as mass spectrometry, Western blotting, ELISA, and more. Higher protein yields are achieved by ExoQuick purification than by chromatography, DynaBeads, or ultracentrifugation.

Exosomes isolated with ExoQuick-TC also provide excellent samples for studying exosome-associated nucleic acids such as microRNAs, siRNAs, and even mRNA. Quantitative analytical techniques such as qPCR, microarray studies, and next-generation sequencing are all compatible with nucleic acids isolated from ExoQuick-TC-purified exosomes.

Backed by a growing number of publications, ExoQuick-TC is often the best option for researchers working with low sample volumes, such as clinical research samples or small animal models.

ExoQuick-TC exosome isolation methods are patented technologies⁴.

Choose the right ExoQuick for your application:

REFERENCES

1. Chugh PE, et al. Systemically Circulating Viral and Tumor-Derived MicroRNAs in KSHV-Associated Malignancies. PLoS Pathog. 2013. 9(7): e1003484. PMCID: PMC3715412.

2. Umezu T, et al. Leukemia cell to endothelial cell communication via exosomal miRNAs. Oncogene. 2013 May 30. 32(22):2747-55. PMID: 22797057.

3. Sohel MM, et al. Exosomal and Non-Exosomal Transport of Extra-Cellular microRNAs in Follicular Fluid: Implications for Bovine Oocyte Developmental Competence. PLoS One. 2013 Nov 4. 8(11):e78505. PMCID: PMC3817212.

4. Antes T, et al. Methods for Microvesicle Isolation and Selective Removal. Patent No.: US 9,005,888 B2.

How It Works

High-throughput, quantitative exosome recovery

ExoQuick-TC can be used to purify exosomes from a wide variety of tissue culture media⁴, and from certain biofluids such as saliva¹, urine², follicular fluid³, and breast milk⁵. With a simple workflow involving minimal hands-on time and low input sample volume requirements, ExoQuick-TC is an excellent option for researchers who need to purify multiple exosome samples.

To isolate exosomes from tissue culture media, simply:

• Add an appropriate volume of ExoQuick-TC
• Incubate overnight at 4°C
• Isolate exosomes with a 30-minute low-speed spin (1500g).

Isolated exosomes can be found in the pellet and resuspended in an appropriate solution.

You can verify the presence of exosomes with a number of different methods, including Western blotting for general exosome markers (CD63, CD9, CD81, and HSP70), NanoSight analysis, or EM (learn about different ways to detect exosomes and more in our Exosome Basics Guide).

The Bottom LineWith ExoQuick-TC, you can obtain high-quality exosomes from tissue culture media and certain biofluids using a protocol that can easily be performed on multiple samples and requires very low volumes of input sample.

REFERENCES

1. Yang J, et al. Detection of Tumor Cell-Specific mRNA and Protein in Exosome-Like Microvesicles from Blood and Saliva. PLoS ONE. 2014. 9(11): e110641. PMCID: PMC 4232306.

2. Alvarez ML. Isolation of urinary exosomes for RNA biomarker discovery using a simple, fast, and highly scalable method. Methods Mol Biol. 2014. 1182:145-70. PMID: 25055908.

3. Sohel MM, et al. Exosomal and Non-Exosomal Transport of Extra-Cellular microRNAs in Follicular Fluid: Implications for Bovine Oocyte Developmental Competence. PLoS One. 2013 Nov 4. 8(11):e78505. PMCID: PMC3817212.

4. Zhu L, et al. Novel method for extracting exosomes of hepatocellular carcinoma cells. World J Gastroenterol. 2014 June 7. 20(21): 6651-6657. PMCID: PMC4047354.

5. Gu Y, et al. Lactation-Related MicroRNA Expression Profiles of Porcine Breast Milk Exosomes. PLoS ONE. 2012. 7(8): e43691. PMCID: PMC3427246.

Supporting Data

Characterizing ExoQuick-TC exosomes with NanoSight

Exosomes purified with ExoQuick-TC from tissue culture media show the expected particle size distribution and high concentration yields when analyzed using NanoSight’s Nanoparticle Tracking Analysis (NTA, Figure 1).

Figure 3.Exosome size distribution and yields from tissue culture media. Cells from a human HT1080 lung sarcoma cell line were cultured in conditioned media (serum-free) for 72 hours. 10 mL of media was combined with 2 mL ExoQuick-TC, incubated overnight at 4°C, and centrifuged at 1500g for 30 minutes to isolate exosomes. The exosome pellet was resuspended in 1 mL PBS, diluted 1:40, and visualized on the NanoSight LM10 instrument. The analysis shows that ExoQuick-TC recovered 133 nm exosomes at a concentration of 1.74 x 10⁹ particles/mL.

Resources

Citations

• Kong, J, et al. (2018) Exosomes of endothelial progenitor cells inhibit neointima formation after carotid artery injury. Journal of Surgical Research.2018 Dec 1; 232:398-407. Link:Journal of Surgical Research
• Yang, X, et al. (2018) Exosome-encapsulated antibiotic against intracellular infections of methicillin-resistant Staphylococcus aureus. Int J Nanomedicine.2018 Nov 29; 13:8095-8104. PM ID:30555228
• Wang, D, et al. (2018) Exosomes from mesenchymal stem cells expressing miR-125b inhibit neointimal hyperplasia via myosin IE. J. Cell. Mol. Med..2018 Nov 28;. PM ID:30484954
• Li, D, et al. (2018) Exosomes Derived From miR-133b-Modified Mesenchymal Stem Cells Promote Recovery After Spinal Cord Injury. Front Neurosci.2018 Nov 22; 12:845. PM ID:30524227
• Naseri, Z, et al. (2018) Exosome-mediated delivery of functionally active miRNA-142-3p inhibitor reduces tumorigenicity of breast cancer in vitro and in vivo. Int J Nanomedicine.2018 Nov 20; 13:7727-7747. PM ID:30538455
• Chen, T, et al. (2018) Increased urinary exosomal microRNAs in children with idiopathic nephrotic syndrome. EBioMedicine.2018 Nov 19;. PM ID:30467011
• Kuroda, H, et al. (2018) Cluster of Differentiation 46 is the Major Receptor in Human Blood-Brain Barrier Endothelial Cells for Uptake of Exosomes Derived from Brain-Metastatic Melanoma Cells (SK-Mel-28). Mol. Pharm..2018 Nov 19;. PM ID:30452273
• Hosaka, T, et al. (2018) ADAR2-dependent A-to-I RNA editing in the extracellular linear and circular RNAs. Neurosci. Res..2018 Nov 15;. PM ID:30448461
• Faict, S, et al. (2018) Exosomes play a role in multiple myeloma bone disease and tumor development by targeting osteoclasts and osteoblasts. Blood Cancer J.2018 Nov 8; 8(11):105. PM ID:30409995
• Sagini, K, et al. (2018) Oncogenic H-Ras Expression Induces Fatty Acid Profile Changes in Human Fibroblasts and Extracellular Vesicles. Int J Mol Sci.2018 Nov 8; 19(11). PM ID:30413053
• Mead, B, Ahmed, Z & Tomarev, S. (2018) Mesenchymal Stem Cell-Derived Small Extracellular Vesicles Promote Neuroprotection in a Genetic DBA/2J Mouse Model of Glaucoma. Invest. Ophthalmol. Vis. Sci..2018 Nov 1; 59(13):5473-5480. PM ID:30452601
• Vracar, TC, et al. (2018) Enoxacin and bis-enoxacin stimulate 4T1 murine breast cancer cells to release extracellular vesicles that inhibit osteoclastogenesis. Sci Rep.2018 Nov 1; 8(1):16182. PM ID:30385810
• Brenner, A, Su, G & Momen-Heravi, F. (2018) Isolation of Extracellular Vesicles for Cancer Diagnosis and Functional Studies. Methods in Molecular Biology.2018 Oct 31;:229-237. Link:Methods in Molecular Biology
• Yamauchi, M, et al. (2018) Efficient method for isolation of exosomes from raw bovine milk. Drug Dev Ind Pharm.2018 Oct 26;:1-18. PM ID:30366501
• Kitagawa, T, et al. (2018) Circulating pancreatic cancer exosomal RNAs for detection of pancreatic cancer. Mol Oncol.2018 Oct 25;. PM ID:30358104
• Wang, M, et al. (2018) The Role of CXCL12 Axis in Lung Metastasis of Colorectal Cancer. J. Cancer.2018 Oct 25; 9(21):3898-3903. Link:J. Cancer
• Frassanito, MA, et al. (2018) Bone marrow fibroblasts overexpress miR-27b and miR-214 in step with multiple myeloma progression, dependent on tumour cell-derived exosomes. J. Pathol..2018 Oct 25;. PM ID:30357841
• Liu, B, et al. (2018) Downregulation of microRNA‑30a in bronchoalveolar lavage fluid from idiopathic pulmonary fibrosis patients. Mol Med Rep.2018 Oct 18;. PM ID:30365083
• Leszczynska, A, et al. (2018) Exosomes from normal and diabetic human corneolimbal keratocytes differentially regulate migration, proliferation and marker expression of limbal epithelial cells. Sci Rep.2018 Oct 11; 8(1):15173. PM ID:30310159
• Momen-Heravi, F & Bala, S. (2018) Extracellular vesicles in oral squamous carcinoma carry oncogenic miRNA profile and reprogram monocytes via NF-κB pathway. Oncotarget.2018 Oct 5; 9(78):34838-34854. PM ID:30410681