EGFR-AF555, EGFRvIII, PD-L1-AF647, and PDGFRα-AF555 antibodies were purchased from Cell Signaling Technology, IDH1 and PD-L2-AF647 antibodies were from BioLegend, and IDH1-R132H antibody was from EMD Millipore. CD63 antibody was acquired from Ancell Corporation. CD9 antibody was acquired from Abcam. CD81 antibody was acquired from Santa Cruz Biotechnology. Vendor and clone information is summarized in Supplementary Table 3 . IDH1, CD63, and CD81 antibodies were conjugated to Alexa Fluor 555 and EGFRvIII, IDH1-R132H, PDPN, and CD9 antibodies were conjugated to Alexa Fluor 647 utilizing the Alexa Fluor 555/647 Labeling Kits per kit instructions (Thermo Fisher Scientific). We used Alexa Fluor 488 for the streptavidin imaging channel and Cy5 channel for the quenching test.
Cd81 antibody
Manufactured by Santa Cruz Biotechnology
Sourced in United States
The CD81 antibody is a laboratory reagent used to detect the presence and expression of the CD81 protein in biological samples. CD81 is a cell surface protein that plays a role in cell signaling and immune function. The antibody can be used in various immunological techniques, such as flow cytometry, immunohistochemistry, and Western blotting, to analyze the expression and distribution of CD81 in different cell types and tissues.
Automatically generated - may contain errors
Lab products found in correlation
Cd63 antibody, by Santa Cruz Biotechnology (2 mentions)
Egfrviii, by Cell Signaling Technology (1 mentions)
Cd9 antibody, by Abcam (1 mentions)
Idh1 r132h, by Thermo Fisher Scientific (1 mentions)
Dexamethasone d1756, by Merck Group (1 mentions)
Pkh26 red fluorescent cell linker kit for general cell membrane labeling, by Merck Group (1 mentions)
Ab108337, by Abcam (1 mentions)
Calnexin, by Cell Signaling Technology (1 mentions)
Pkh67 green fluorescent cell linker kit for general cell membrane labeling, by Merck Group (1 mentions)
Fitc conjugated goat anti rabbit igg h l secondary antibody, by Thermo Fisher Scientific (1 mentions)
Grp94, by Cell Signaling Technology (1 mentions)
Lamin a c, by Cell Signaling Technology (1 mentions)
Pierce bca protein kit, by Thermo Fisher Scientific (1 mentions)
Tsg101 sc 7964, by Santa Cruz Biotechnology (1 mentions)
Hrp conjugated antibodies, by Agilent Technologies (1 mentions)
Ripa lysis buffer, by Beyotime (1 mentions)
Ecl star, by Beyotime (1 mentions)
Cd9 antibody, by Santa Cruz Biotechnology (1 mentions)
4 protocols using cd81 antibody
1
Multicolor Antibody Labeling for Flow Cytometry
2
Exosome Isolation and Membrane Labeling
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Exosome isolation reagent (from cell culture media) was purchased from Thermo Fisher scientific (4478359, China). PKH67 Green Fluorescent Cell Linker Kit for General Cell Membrane Labeling (MINI67-1KT, China) and PKH26 Red Fluorescent Cell Linker Kit for General Cell Membrane Labeling (PKH26GL-1KT, China) were purchased from Sigma. Lipopolysaccharide (LPS, Escherichia coli 0111: B4, L4391), β‐glycerophosphate (G9422), L‐ascorbic acid 2‐phosphate (49752), and dexamethasone (D1756) were from Sigma, China. Antibodies used in this study were rabbit polyclonal antibody against alkaline phosphatase (ALP) (ab108337, Abcam), CD63 Antibody (sc-5275, SANTA CRUZ Biotechnology), CD81 Antibody (sc-166029, SANTA CRUZ Biotechnology), calnexin ( 2679 T, cell Signaling Technology), Grp94 (20292 T, cell Signaling Technology), and Lamin A/C (4777 T, cell Signaling Technology). Secondary antibody used for immunofluorescence staining was fluorescein isothiocyanate (FITC)-conjugated goat anti-rabbit IgG (H + L) Secondary Antibody (31635, Thermo Fisher Scientific, China).
3
Western Blot Analysis of EV Proteins
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The protein-based concentration of EVs, CM and Sup was measured using Pierce BCA Protein Kit (Cat # 23225, ThermoFisher Scientific) for bicinchoninic acid assay according to the protocols provided by the manufacturer. Samples (40 µg proteins) were separated on 12.5% sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) by applying 120 Volt (V) in running buffer for 90 min. In the next step, proteins were transferred to the nitrocellulose membranes by applying 110 V in transfer buffer for 70 min. Membrane blocking was done with 5% skimmed milk prepared in Tris-buffered saline containing 0.1% Tween 20 (1 × TBST). Incubation of membranes with primary antibodies was performed overnight at 4 °C. Primary antibodies included: TSG101 (SC-7964, Santa Cruz Biotechnology) with 1:200 dilution, CD81 Antibody (SC-166029, Santa Cruz Biotechnology) with 1:200 dilution and CD9 (Ab92726, Abcam) with 1:1000 dilution. Afterwards, membranes were incubated for 1 h with respective secondary antibodies (HRP conjugated antibodies) (Dako, Agilent Denmark).
4
Exosome Protein Profiling by Western Blot
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After isolating the protein with radioimmunoprecipitaion assay (RIPA) lysis buffer (Beyotime), separation with a 4-20% precast gel (Willget, Shanghai, China) was completed. The protein was then transferred to a nitrocellulose membrane (PALL, New York, NY, USA). Then, the sections were incubated with CD9 antibody (1:1,000, Santa Cruz, Santa Cruz City, CA, USA), CD63 antibody (1:500, Santa Cruz) and CD81 antibody (1:500, Santa Cruz) at 4℃ overnight. After washing, the membrane was incubated with goat anti-mouse IgG (H+L) secondary antibody (1:20,000, Santa Cruz) for 2 hours (h). Finally, the protein band was visualized with ECL Star (Beyotime).
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