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Exoquick tc exosome precipitation solution kit

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ExoQuick-TC Exosome Precipitation Solution Kit is a laboratory product designed to isolate and concentrate exosomes from cell culture media or other biological fluids. The kit utilizes a proprietary precipitation reagent to selectively isolate extracellular vesicles, including exosomes, from the sample.

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Lab products found in correlation

Amicon ultra 15 centrifugal filter unit, by Merck Group (1 mentions) Nah2po4, by Merck Group (1 mentions) Bca kit, by Beyotime (1 mentions) Tecnai 10, by Philips (1 mentions) Nta software, by Malvern Panalytical (1 mentions) Ultracentrifuge, by Beckman Coulter (1 mentions) Nanoplus, by Beckman Coulter (1 mentions) Bicinchoninic acid bca kit, by Beyotime (1 mentions) Amicon ultra 15, by Merck Group (1 mentions)

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ExoQuick-TC (231 citations) ExoQuick-TC kit (67 citations) ExoQuick solution (48 citations) ExoQuick precipitation kit (33 citations) ExoQuick precipitation solution (26 citations) ExoQuick-TC solution (18 citations) ExoQuick-TCTM (12 citations) ExoQuick-TC Precipitation Solution (11 citations) Exosome precipitation kit (9 citations) ExoQuick exosome precipitation (5 citations)

12 protocols using exoquick tc exosome precipitation solution kit

1

Exosome Isolation from HUVECs with Phosphate Modulation

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HUVECs were cultured in DMEM with the addition of NaH2PO4 for the HP (3 mM; pH=7.4) group or without NaH2PO4 for the no phosphorus (NP) group for 48 h at 37°C in a humidified atmosphere with 5% CO2 for exosome isolation. NaH2PO4 (MilliporeSigma) was used to increase the concentration of phosphorus in the medium. Before extraction of exosomes from HUVECs, the cells were cultured with medium supplemented with 10% exosome-depleted FBS for 48 h at 37°C with 5% CO2 (C38010100; Shanghai VivaCell Biosciences, Ltd.).
Exosomes were concentrated using an ExoQuick-TC Exosome Precipitation Solution kit (EXOTC10A; System Biosciences, LLC). This method has been widely used before and has been proven to collect exosomes effectively (21 (link)-23 (link)). Briefly, the supernatant was centrifuged at room temperature as follows: 300 × g for 10 min, 2,000 × g for 30 min and 10,000 × g for 30 min. An Amicon Ultra15 Centrifugal Filter Unit (100 kDa; MilliporeSigma) was used to concentrate the supernatant. The ultrafiltration liquid and exosome isolation reagents were mixed at a 5:1 ratio and incubated at 4°C for ~16 h. Finally, the mixture was centrifuged at room temperature at 1,500 × g for 30 min, and the exosome pellets were resuspended in 200 µl PBS. Protein quantification of exosomes was performed using a BCA kit (Beyotime Institute of Biotechnology).
Qin Z., Li Y., Li J., Jiang L., Zhang Z., Chang K., Yang Q., Chen S., Liao R, & Su B. (2022). Exosomal STAT1 derived from high phosphorus-stimulated vascular endothelial cells induces vascular smooth muscle cell calcification via the Wnt/β-catenin signaling pathway. International Journal of Molecular Medicine, 50(6), 139.
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2

Urine Exosome Isolation Workflow

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Morning urine of the patients was processed within 4 h after voiding, as no degradation was observed within this period in pre-trials. Approximately 50–100 mL of urine was transferred to tubes and centrifuged for 10 min at 2000× g to remove any cells or cell debris. The supernatant was transferred to fresh tubes and stored at −80 °C or processed to isolate exosomes within a week. Exosomes from the urine were isolated by using an ExoQuick-TC Exosome Precipitation Solution Kit (EXOTC10A-1, System Biosciences, Berkeley, CA, USA). In brief, 250 μL ExoQuick solution was added to 1 mL of concentrated urine (1:4), mixed thoroughly, and incubated overnight at 4 °C. Then the mixture was centrifuged at 1500× g for 30 min at 4 °C, and the supernatant was removed. The exosome pellet was resuspended in 200 μL of PBS.
Umair Z., Baek M.O., Song J., An S., Chon S.J, & Yoon M.S. (2022). MicroRNA-4516 in Urinary Exosomes as a Biomarker of Premature Ovarian Insufficiency. Cells, 11(18), 2797.
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3

Exosome Isolation from Mouse Brain Endothelial Cells

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Mouse brain ECs were cultured as described above. Exosomes were isolated using ExoQuick-TC (Exosome Precipitation Solution kit, System Biosciences) following previously described protocol12 (link), and suspended in PBS. EC-Exo are from non-diabetic condition. Briefly, media was isolated from a 100% confluent T75 tissue culture flask (containing ~5–6 million cells) containing 10 ml of media with exosome free FBS. This media was centrifuged and filtered via a 0.22 μm syringe filter before adding ExoQuickTC (2 ml for 10 ml media). Exosome number was counted using the IZON qNano device (Izon). To decrease the heterogeneity of Exo treatments, we tightly controlled the cell culture conditions, such as passage, density, and culture time. Liposomes are synthetic versions of exosomes mimicking the exosomal lipid layer and were generated using thin-film hydration technique as previously described13 (link).
Venkat P., Cui C., Chopp M., Zacharek A., Wang F., Landschoot-Ward J., Shen Y, & Chen J. (2019). MiR-126 mediates brain endothelial cell exosome treatment induced neurorestorative effects after stroke in T2DM mice. Stroke, 50(10), 2865-2874.
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4

Exosome Isolation from Follicular Fluid and Cell Culture

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The follicular fluid exosomes were collected by ExoQuick‐TC Exosome Precipitation Solution Kit (System Biosciences). In brief, 250 µL ExoQuick Solution was added to the 1 mL follicular fluid (1:4), mixed well and incubated overnight at 4°C. Then, the mixture was centrifuged at 1500 g for 30 minutes at 4°C, and the supernatant was removed. The exosome pellet was resuspended in 500 µL PBS and passed through a 0.22‐µm filter.
Exosomes from cell culture supernatants were isolated by ultracentrifugation. In brief, 293T cells were starved in medium containing 1% BSA for 48 hours. Then, cell supernatant was centrifuged at 300 g for 10 minutes to remove cells. The supernatant fluid was then centrifuged at 2000 g for 10 minutes at 4°C to remove dead cells. The resultant supernatant fluid was transferred to an ultracentrifuge tube and centrifuged at 100 000 g for 2 hours. The pellet was suspended in PBS and filtered through a 0.22‐μm filter, and then centrifuged at 100 000 g for 2 hours. The pellet was resuspended in 200 μL PBS and stored at −80°C.
Li H., Huang X., Chang X., Yao J., He Q., Shen Z., Ji Y, & Wang K. (2019). S100‐A9 protein in exosomes derived from follicular fluid promotes inflammation via activation of NF‐κB pathway in polycystic ovary syndrome. Journal of Cellular and Molecular Medicine, 24(1), 114-125.
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5

Exosome Isolation from NSCLC Cell Cultures

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The NSCLC cell lines were cultured in DMEM containing 10% exosome-depleted FBS. After 48 h, the medium was collected for exosome isolation. Briefly, the culture medium was collected into an ultrafiltration tube by centrifugation at 2000×g for 10 min. To purify exosomes from the medium, we used the ExoQuick-TC™ Exosome Precipitation Solution Kit (System Biosciences, Palo Alto, CA, USA) according to the manufacturer’s instructions. Subsequently, the exosome pellets were resuspended in PBS and filtered (0.22-μm pore size; Millipore, Billerica, MA, USA) for functional assays or protein detection. The size and quality of the exosomes were determined using an LM10-HS NanoSight instrument (Malvern Instruments, Malvern, UK) and NTA software (Malvern Instruments). The exosomes were examined using a transmission electron microscope (PHILIPS-TECNAI 10, Philips, Amsterdam, Netherlands) at 120 kV. For in vitro assays, purified exosomes were used at 20 µg/ml.
Song J.W., Zhu J., Wu X.X., Tu T., Huang J.Q., Chen G.Z., Liang L.Y., Zhou C.H., Xu X, & Gong L.Y. (2021). GOLPH3/CKAP4 promotes metastasis and tumorigenicity by enhancing the secretion of exosomal WNT3A in non-small-cell lung cancer. Cell Death & Disease, 12(11), 976.
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6

Isolation of Follicular Fluid Exosomes

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Exosomes from follicular fluid were isolated by ExoQuick-TC Exosome Precipitation Solution Kit (System Biosciences). In brief, 250μL ExoQuick Solution was added to the 1 mL follicular fluid (1:4), mixed well and incubated overnight at 4°C. Then, the mixture was centrifuged at 1,500 g for 30 minutes at 4°C, and the supernatant was removed. The exosome pellet was resuspended in 500μL PBS and passed through a 0.22- μm filter.
Huang X., Wu B., Chen M., Hong L., Kong P., Wei Z, & Teng X. (2020). Depletion of exosomal circLDLR in follicle fluid derepresses miR-1294 function and inhibits estradiol production via CYP19A1 in polycystic ovary syndrome. Aging (Albany NY), 12(15), 15414-15435.
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7

Isolation and Characterization of Renal EVs

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Renal primary epithelial cells and macrophage-derived EVs isolated from T2D mice were collected over a 24 h time period in EV-depleted complete medium, which was prepared by overnight centrifugation at 100,000 × g at 4 °C. Unless stated otherwise, EVs were isolated from cell culture medium by differential ultracentrifugation using a modified version of a protocol by Thery et al. 17 . The collected medium was depleted of cells and cell debris by consecutive, low-speed centrifugation (2,000 × g for 15 min and 16,000 × g for 20 min). The supernatants obtained were carefully collected and centrifuged for 90 min at 100,000 × g at 4 °C. Pellets from this centrifugation step were washed in PBS, pooled, and centrifuged again for 90 min at 100,000 × g at 4 °C. The obtained pellets were resuspended in lysis buffer (see below), PBS solution or RPMI-1640 medium, depending on subsequent experiments. EV solutions intended for cell treatment were sterile filtered through a 0.22 µm syringe filter. Resuspended EVs were either used for subsequent analysis or aliquoted and stored at -80 °C. For isolation of EVs derived from primary cells, a commercially available ExoQuick-TC exosome precipitation solution kit from System Biosciences (Palo Alto, CA) was utilized.
Jiang W.J., Xu C.T., Du C.L., Dong J.H., Xu S.B., Hu B.F., Feng R., Zang D.D., Meng X.M., Huang C., Li J, & Ma T.T. (2022). Tubular epithelial cell-to-macrophage communication forms a negative feedback loop via extracellular vesicle transfer to promote renal inflammation and apoptosis in diabetic nephropathy. Theranostics, 12(1), 324-339.
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8

Isolation of Follicular Fluid Exosomes

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All of the patients received controlled ovarian hyperstimulation by using a combination of gonadotropin-releasing hormone (GnRH) agonist or antagonist and recombinant follicle-stimulating hormone (FSH). FF was collected by using transvaginal ultrasound-guided aspiration from dominant follicles (> 18 mm), (yielding 2–3 ml) at 34 to 36 h after the administration of 10,000 IU human chorionic gonadotropin (hCG). The supernatants were stored at − 80 °C after centrifugation at 3000 ×g for 15 min to remove cell debris and other particles. The FF exosomes were collected with an ExoQuick-TC Exosome Precipitation Solution Kit (System Biosciences, Palo Alto, CA), which is widely used to obtain exosomes from different body fluids. However, the obtained exosome sample was not homogenous. In brief, 250 µl of ExoQuick Solution was added to 1 ml of FF in a 1:4 ratio. The mixture was thoroughly mixed and left to incubated overnight at a temperature of 4 °C. After incubation, the mixture was centrifuged at 1500 ×g for 30 min at 4 °C and the supernatant was carefully removed. The resulting exosome pellet was then resuspended in 500 µl of PBS and passed through a 0.22 µm filter to obtain the desired solution.
Gu Y., Zhang X., Wang R., Wei Y., Peng H., Wang K., Li H, & Ji Y. (2024). Metabolomic profiling of exosomes reveals age-related changes in ovarian follicular fluid. European Journal of Medical Research, 29, 4.
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9

Isolation and Characterization of Breast Cancer-Derived Exosomes

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MCF-7 cells were placed in serum-free culture medium for isolation of exosomes from BC cell-derived conditioned medium. Cell supernatant was centrifuged for 15 min at 300 g to remove cells and for 30 min at 3000 g to remove cell debris. Then, the supernatant was centrifuged on an ultracentrifuge (Beckman, Brea, CA, USA) for 2 h at 100 000 g to sediment exosomes. The supernatant was added with the ExoQuick-TC Exosome Precipitation Solution kit (System Biosciences, Johnstown, PA) for exosome isolation. Afterward, isolated exosomes were suspended in PBS and stored at −80°C.
The isolated exosomes were diluted to 500 ng/ml and analyzed using Nanoplus (Beckman Coulter, Miami, FL, USA) for particle size analysis. Exosomes were loaded to a copper mesh, stained with 2% uranyl acetate and dried. Exosome morphology was photographed using a transmission electron microscope (JEM-2000EX, Tokyo, Japan), and the expression of exosome markers CD81 and CD63 as well as a Golgi matrix protein GM130 was investigated by western blotting.
Sun S., Wang Z., Yao F., Sun K., Li Z., Sun S, & Li C. (2023). Breast cancer cell-derived exosome-delivered microRNA-155 targets UBQLN1 in adipocytes and facilitates cancer cachexia-related fat loss. Human Molecular Genetics, 32(13), 2219-2228.
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10

Exosome Isolation from ESRD, RTR, and Healthy Plasma

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Blood was obtained from patients with ESRD, RTR, and Nor, respectively. The blood was centrifuged at 3,000 × g for 20 min, and the supernatant plasma was subjected to the ExoQuick-TC Exosome Precipitation Solution Kit (System Biosciences, USA) for exosomes extraction. Briefly, 250 μl of plasma was added to 63 μl ExoQuick Exosomes Precipitation Reagent and fully mixed. After the mixture had been incubated at 4°C for 30 min, it was centrifuged at 1,500 × g for 30 min at room temperature (RT), and the yellow or white precipitate at the bottom of the EP tube was the exosomes. Then, the supernatant was removed by aspiration, and the exosomes pellets were centrifuged again at 1,500 × g for 5 min. Finally, the exosomes pellets were resuspended in 200 μl phosphate-buffered saline (PBS). The protein content of exosomes was determined by the bicinchoninic acid (BCA) kit (Beyotime Biotechnology, Shanghai, China).
Lin X., Zhu T., Xu F., Zhong J.Y., Li F., Shan S.K., Wu F., Guo B., Zheng M.H., Wang Y., Xu Q.S., Liao X.B., Lu H.Y., Xie X.B, & Yuan L.Q. (2021). Plasma Exosomes Derived From Patients With End-Stage Renal Disease and Renal Transplant Recipients Have Different Effects on Vascular Calcification. Frontiers in Cell and Developmental Biology, 8, 618228.
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