Journal: PLOS ONE

Article Title: Use of extracellular vesicle microRNA profiles in patients with acute myeloid leukemia for the identification of novel biomarkers

doi: 10.1371/journal.pone.0306962

Figure Lengend Snippet: (A) The size and concentration trend of vesicles by fractions are presented. The eluted fractions (11 and 12; 0.5 mL each) were used for vesicle isolation. Samples were diluted 10-fold. (B) The size distribution of the isolated vesicles was determined using nanoparticle tracking analysis (NTA). The average size of BM aspirate serum or plasma-derived vesicles was 115.5 ± 2.7 nm and 101.5 ± 4.4 nm, respectively; these sizes were within the size range of typical EVs. Samples were diluted 10-fold. (C) In transmission electron microscopy (TEM) images, the size of isolated vesicles was <200 nm, and they were visualized as cup-shaped vesicles under high magnification. (D) Western blotting showed that the isolated vesicles were positive for the markers of EVs (CD63 and CD81). EVs, extracellular vesicles; BM, bone marrow; AML, acute myelogenous leukemia.

Article Snippet: After blocking with 5% skim milk (w/v) in 0.1% TBST for 2 h, the membranes were probed overnight at 4°C with 1:1000 dilutions of rabbit anti-CD63 polyclonal antibody and rabbit anti-CD81 polyclonal antibodies (all from Bioss Antibodies, MA, USA).

Techniques: Concentration Assay, Isolation, Derivative Assay, Transmission Assay, Electron Microscopy, Western Blot

Journal: bioRxiv

Article Title: Immunojanus Particles for low-volume and isolation-free unlabeled characterization of small Extracellular Vesicle in biofluids: Characterization of disease type by surface marker profiling

doi: 10.1101/2024.08.17.607528

Figure Lengend Snippet: (a) Characterization of CD63+, CD81+, GPC1+, CEA+, and aEGFR+ sEVs in cell media (DiFi, GBM6, MDA-MB-468, A375P, 3T3) and human biofluids (urine and serum) using IJPs. (b) Characterization of the same sEVs using UC + SPR, showing identical results. Panels (c-e) show immunostaining of IJPs (red) with anti-CD63 capture mixed with samples containing Quantum Dots labeled with anti-CD63 (green): (c) PBS, (d) 1E8 sEVs/mL, and (e) 1E10 sEVs/mL. (f) BCA assay of IJPs mixed with 10x diluted human plasma, filtered with 220nm, and washed with PBS, showing high non-specific binding of proteins for controls that do not change the blinking rate. (g) Instead of the BCA assay, anti-CD63 with HRP is added and washed, followed by an enzymatic reaction with TMB. IJPs producing significant shifts in blinking are highly enriched in anti-CD63, while those that do not change blinking significantly do not produce a high signal in this assay, despite having similar total prot ein concentrations as shown in (f). (h) UC + SPR and IJPs show a linear trend for the relative expression of different sEVs normalized by CD63+ sEVs. Error bars represent one standard deviation.

Article Snippet: In this research, the antibodies utilized included CD63 Mouse Monoclonal Antibody (Proteintech, Catalog: 67605-1-Ig, Lot: 10023876), CD81 Rabbit Polyclonal Antibody (Proteintech, Catalog: 27855-1-AP, Lot: 00107630), Glypican 1 Rabbit Polyclonal Antibody (Proteintech, Catalog: 16700-1-AP, Lot: 00056068), CEA Rabbit Polyclonal Antibody (Proteintech, Catalog: 10421-1-AP, Lot: 00017390), Mouse IgG1 Isotype Control Mouse Monoclonal Antibody (Proteintech, Catalog: 66360-1-Ig, Lot: 10028151) and Phospho-Tau181 Monoclonal Antibody (Invitrogen, Reference: MN1050, Lots: XF3582141, YI4023267).

Techniques: Immunostaining, Labeling, BIA-KA, Clinical Proteomics, Binding Assay, Expressing, Standard Deviation

Journal: Journal of Cancer Research and Clinical Oncology

Article Title: Effect of human bone marrow mesenchymal stem cell-derived microvesicles on the apoptosis of the multiple myeloma cell line U266

doi: 10.1007/s00432-024-05822-2

Figure Lengend Snippet: Western blotting assay for the verification of the BMSC-MVs. CD63, CD73, and CD81 with approximately 50, 75, and 26 kDa protein bands, respectively, were detected

Article Snippet: Next, the membranes were separately incubated with the following primary antibodies: rabbit polyclonal anti-human CD63 antibodies (cat. No. 11,271-T16; Sino Biological, China), rabbit monoclonal anti-human CD73 antibodies (cat No. ab124725; Abcam, MA, United States), and rabbit polyclonal anti-human CD81 antibodies (cat No. ab155760; Abcam, MA, United States) as per the suppliers’ instructions.

Techniques: Western Blot