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Invitrogen total exosome isolation kit

Manufactured by Thermo Fisher Scientific
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The Invitrogen Total Exosome Isolation Kit is a laboratory product designed to isolate and purify exosomes from various sample types, including cell culture media and biological fluids. The kit utilizes a proprietary precipitation-based method to separate exosomes from other cellular components, allowing for their subsequent analysis and characterization.

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Lab products found in correlation

Exoquick exosome precipitation solution, by System Biosciences (2 mentions) Bca protein assay kit, by Thermo Fisher Scientific (2 mentions) Invitrogen total exosome isolation reagent, by Thermo Fisher Scientific (1 mentions) All in one mirna qrt pcr kit, by GeneCopoeia (1 mentions) Nucleozol reagent, by Macherey-Nagel (1 mentions) Ns300, by Malvern Panalytical (1 mentions) Jem 1400 electron microscope, by JEOL (1 mentions) M per reagent, by Thermo Fisher Scientific (1 mentions) Exoquick exoq5tm 1, by System Biosciences (1 mentions) Exorneasy serum plasma maxi kit, by Yeasen (1 mentions) C elegans cel mir 39 standard rna, by Sangon (1 mentions) Pierce bicinchoninic bca protein assay, by Thermo Fisher Scientific (1 mentions) Nanosight ns300, by Malvern Panalytical (1 mentions)

6 protocols using invitrogen total exosome isolation kit

1

Exosome Isolation from Murine Samples

Cited 1 time
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At the end of the studies, mice were euthanized with isoflurane and CO2 inhalation and blood samples were withdrawn from the left femoral artery. Exosomes were isolated from blood samples using Invitrogen Total Exosome Isolation Kit (from plasma) (Invitrogen 4484450, Thermo Fisher Scientific, Waltham, MA, USA). For the isolation of exosomes from cell cultures, Invitrogen Total Exosome Isolation Reagent (from cell culture media) (Invitrogen 4478359, Thermo Fisher Scientific, Waltham, MA, USA) was used. All of the reagents were provided by the kits, and procedures adhered to the manufacturers’ instructions. The use of exosome isolation kits and protocols was referred to relevant studies [21 (link),22 (link)].
Wang J.D., Wang Y.Y., Lin S.Y., Chang C.Y., Li J.R., Huang S.W., Chen W.Y., Liao S.L, & Chen C.J. (2021). Exosomal HMGB1 Promoted Cancer Malignancy. Cancers, 13(4), 877.
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2

Exosomal miRNA Isolation and Quantification

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Plasma and serum exosomes were isolated using the Invitrogen™ Total Exosome Isolation Kit (Invitrogen, CA, USA) and ExoQuick™ exosome precipitation solution (System Biosciences, CA, USA), respectively. The extracted exosomes were examined by the JEM-1400 electron microscope (JEOL, Tokyo, Japan), NanoSight NS300 (Malvern, Cambridge, UK), and Western blot analysis with markers as previously reported [15 (link)]. Small RNAs were extracted from the exosomes using the NucleoZOL reagent (Macherey-Nagel, Duren, Germany). Commercial external control for miRNA (TIANGEN BIOTECH, Beijing, China) was used as a reference and detection by the corresponding primer. Exosomal miRNAs were reverse transcribed and detected by qRT-PCR using the All-in-one™ miRNA qRT-PCR Kit (GeneCopoeia, Maryland, VA, USA). The relative expression of exosomal miRNA was estimated by the 2−ΔΔCt method (ΔCt = CtmiRNA-Ctexternal control).
Wang Y., Chen X., Lin H., Sun X., Fong W.P., Wu X., Pan Z., Yuan Y., Liang J., Wang D., Du Z., Xing B, & Li Y. (2021). A Prehepatectomy Circulating Exosomal microRNA Signature Predicts the Prognosis and Adjuvant Chemotherapeutic Benefits in Colorectal Liver Metastasis. Cancers, 13(17), 4258.
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3

Exosome Isolation and Purification Methods

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Exosomes were isolated and purified using ExoQuick (EXOQ5TM-1, System Biosciences, Palo Alto, CA, USA), Invitrogen Total Exosome Isolation Kit (4484451, Thermo Fisher Scientific, Massachusetts, Waltham, USA), and Exosome-Human CD81 Flow Detection Reagent (10622D, Thermo Fisher Scientific, USA) according to the manufacturer’s instructions. Briefly, the reagents were added to CCM or plasma of healthy donors and cancer patients to isolate exosomes and the mixture was vortexed and centrifuged at 4 °C as described in the manufacturers’ protocols. The pellet containing exosomes was resuspended in DPBS or ultrapure water. Subsequently, the exosome pellet was diluted in M-PER reagent (Thermo Fisher Scientific, Massachusetts, Waltham, USA) and BCA reagent A and B (A:B = 50:1) was added and incubated for 30 min at 37 °C. The protein concentration of the pellet was determined using the BCA protein assay kit (Thermo Scientific, Waltham, MA, USA) according to the manufacturer’s instruction. To improve reproducibility, all assays were carried out under same experimental conditions.
Lim J., Choi M., Lee H., Kim Y.H., Han J.Y., Lee E.S, & Cho Y. (2019). Direct isolation and characterization of circulating exosomes from biological samples using magnetic nanowires. Journal of Nanobiotechnology, 17, 1.
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4

Exosome Isolation and RNA Extraction

Cited 2 times
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Exosomes from human and mouse plasma were extracted using an Invitrogen™ Total Exosome Isolation Kit (ThermoFisher, USA) [27 (link)]. Briefly, 200 uL of plasma was mixed with the precipitation reagent provided by the kit and exosomes were isolated according to the manufacturer’s instructions. The exoRNeasy Serum/Plasma Maxi Kit (Yeasen Biotech, Shanghai, China) [28 (link)] was employed to isolate exosomal RNAs in samples previously spiked with 25 fmol of C. elegans cel-miR-39 standard RNA (Sangon, Shanghai, China), used as control. The exosomes were stored at -80° C before cell culture experiments and molecular analyses.
Cao W., Zeng Z., He Z, & Lei S. (2021). Hypoxic pancreatic stellate cell-derived exosomal mirnas promote proliferation and invasion of pancreatic cancer through the PTEN/AKT pathway. Aging (Albany NY), 13(5), 7120-7132.
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5

Plasma Exosome Isolation and Characterization

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Plasma exosomes were isolated using the Invitrogen Total Exosome Isolation Kit (Thermo Fisher Scientific, Waltham, MA, USA) following manufacturer’s instructions. Plasma exosomes were quantitated with measurement of total protein concentration using the Pierce bicinchoninic (BCA) protein assay (Thermo Fisher Scientific, Waltham, MA, USA) following manufacturer’s instructions. Plasma exosomes were then aliquoted into the following exosomal protein amounts for use: 50μg, 100μg, and 200μg.
The size and concentration of exosomes for each sample were evaluated with the Malvern NanoSight NS300 with nanoparticle tracking analysis (NTA) (Malvern Panalytical, Worcestershire, United Kingdom). The suspension of plasma exosomes from each sample was diluted 1:1000 in filtered PBS to obtain 20–100 particles per field. Next, 1mL of the diluted exosome suspension was drawn up and transferred to an O-ring top plate NTA chamber carefully to avoid the introduction of air bubbles. Particles were analyzed using a 488nm laser captured for 60 seconds per run for five total runs per sample; size and concentration of exosome particles were averaged from these five runs.
Munley J.A., Kelly L.S., Gillies G.S., Kannan K.B., Pons E.E., Bible L.E., Efron P.A, & Mohr A.M. (2023). Effects of Trauma Plasma-Derived Exosomes on Hematopoietic Progenitor Cells. Shock (Augusta, Ga.), 59(4), 591-598.
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6

Exosome Isolation Protocol from Cell Culture

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Exosomes were isolated from the cell culture medium using the Invitrogen™ Total Exosome Isolation Kit (ThermoFisher, USA). Specifically, 1 mL of the cell culture medium was combined with the precipitation reagent supplied by the kit, and the exosomes were subsequently isolated following the guidelines provided by the manufacturer. Following this, transmission electron microscopy and western blotting techniques were employed to confirm the morphology and marker proteins of the extracted exosomes. The exosomes were preserved at a temperature of −80°C prior to conducting cell culture experiments and molecular analyses.
Pan F., Pan R., Hu R., Zhang H., Lei S., Zhang L., Zhou C., Zeng Z., Tian X, & Xie Q. (2024). Analysis of the effects of M2 macrophage‐derived PDE4C on the prognosis, metastasis and immunotherapy benefit of osteosarcoma. Journal of Cellular and Molecular Medicine, 28(10), e18395.
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