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$Mitochondrial components segregate into MVBs and are secreted in exosomes. a Confocal co-localization analysis in HEK293 cells treated with FCCP (4 h) and stained with SSBP1 (red), EEA1 and CD63 (green). Bar, 10 µm. Graph shows means for Mander’s co-localization coefficient for SSBP1 and EEA1 or CD63 ( n = 3). t -test: *** P -value < 0.0001. b Confocal co-localization analysis in HEK293 cells co-transfected with Rab7-Q67L-GFP (Rab7mutGFP, green) and mitoDsRed (red), and immunostained for TFAM (purple) and nuclei (HOECHST 58, blue). Center and right images show high magnification of left images (Rab7-Q67L-GFP and mitoDsRed in upper panels; Rab7-Q67L-GFP and TFAM in lower panels). Charts: Fluorescence profiles along the corresponding white lines. Bar, 10 µm. c Mitochondrial components in the exosome fraction obtained from equal numbers of shControl and shnSMase2 J77 T cells. ATAD3, TFAM, and CD63 were detected by immunoblot; mtDNA was detected by PCR for HVRII . Graph: Quantification of exosomal proteins and mtDNA in a representative experiment ( n = 3). d Flow cytometry analysis of mitochondrial mass (Mitotracker green), intracellular ROS levels (DCFDA staining), oxidized DNA (8-OHdG Ab staining), and endogenous TFAM in HEK293 cells knocked down for nSMase2 and Rab27a. Graphs: Quantification from 5–8 independent experiments (Mean). t -test * P -value < 0.05; ** P -value < 0.001. e Electron microscopy images show defects in mitochondrial ultrastructure and cristae organization in shnSMase2 and <t>shRab27a</t> HEK293 cells. Graph: Quantification of mitochondrial cristae width (Mean). t -test: *** P -value < 0.0001. f Graph: Basal oxygen consumption rate (OCR) of control, shnSMase2 and shRab27a HEK293 cells. Dots represent mean from three independent experiments run in duplicate or triplicate. t -test, * P -value < 0.05, *** P -value < 0.0001. Chart: OCR from shControl, shnSMase2 and shRab27a HEK293 cells in response to oligomycin (Oligo), fccp, and rotenone plus antimycin A (Rot/AA). ( n = 2; mean ± S.E.M.). g Western blot analysis of exosomes from Jurkat T cells left untreated, serum-starved overnight or treated with bafilomycin A. Membranes were blotted for TFAM, CD81, and CD63. Graph: Nanoparticle concentration in the exosomal fractions (mean, two independent experiments). h Western blot analysis of exosomes obtained from Jurkat T cells transfected with control or LC3 siRNA. Graph: Nanoparticle concentrations in the exosomal fractions (mean, n = 2). Western blots are representative out of three independent experiments$
Shrab27a Shrnas, supplied by Addgene inc, used in various techniques. Bioz Stars score: 93/100, based on 4 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/product/shrab27a+shrnas/pmc06037695-306-56-43?v=Addgene+inc
Average 93 stars, based on 4 article reviews

shrab27a shrnas - by Bioz Stars, 2026-07

93/100 stars

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1) Product Images from "Priming of dendritic cells by DNA-containing extracellular vesicles from activated T cells through antigen-driven contacts"

Article Title: Priming of dendritic cells by DNA-containing extracellular vesicles from activated T cells through antigen-driven contacts

Journal: Nature Communications

doi: 10.1038/s41467-018-05077-9

$Mitochondrial components segregate into MVBs and are secreted in exosomes. a Confocal co-localization analysis in HEK293 cells treated with FCCP (4 h) and stained with SSBP1 (red), EEA1 and CD63 (green). Bar, 10 µm. Graph shows means for Mander’s co-localization coefficient for SSBP1 and EEA1 or CD63 ( n = 3). t -test: *** P -value < 0.0001. b Confocal co-localization analysis in HEK293 cells co-transfected with Rab7-Q67L-GFP (Rab7mutGFP, green) and mitoDsRed (red), and immunostained for TFAM (purple) and nuclei (HOECHST 58, blue). Center and right images show high magnification of left images (Rab7-Q67L-GFP and mitoDsRed in upper panels; Rab7-Q67L-GFP and TFAM in lower panels). Charts: Fluorescence profiles along the corresponding white lines. Bar, 10 µm. c Mitochondrial components in the exosome fraction obtained from equal numbers of shControl and shnSMase2 J77 T cells. ATAD3, TFAM, and CD63 were detected by immunoblot; mtDNA was detected by PCR for HVRII . Graph: Quantification of exosomal proteins and mtDNA in a representative experiment ( n = 3). d Flow cytometry analysis of mitochondrial mass (Mitotracker green), intracellular ROS levels (DCFDA staining), oxidized DNA (8-OHdG Ab staining), and endogenous TFAM in HEK293 cells knocked down for nSMase2 and Rab27a. Graphs: Quantification from 5–8 independent experiments (Mean). t -test * P -value < 0.05; ** P -value < 0.001. e Electron microscopy images show defects in mitochondrial ultrastructure and cristae organization in shnSMase2 and shRab27a HEK293 cells. Graph: Quantification of mitochondrial cristae width (Mean). t -test: *** P -value < 0.0001. f Graph: Basal oxygen consumption rate (OCR) of control, shnSMase2 and shRab27a HEK293 cells. Dots represent mean from three independent experiments run in duplicate or triplicate. t -test, * P -value < 0.05, *** P -value < 0.0001. Chart: OCR from shControl, shnSMase2 and shRab27a HEK293 cells in response to oligomycin (Oligo), fccp, and rotenone plus antimycin A (Rot/AA). ( n = 2; mean ± S.E.M.). g Western blot analysis of exosomes from Jurkat T cells left untreated, serum-starved overnight or treated with bafilomycin A. Membranes were blotted for TFAM, CD81, and CD63. Graph: Nanoparticle concentration in the exosomal fractions (mean, two independent experiments). h Western blot analysis of exosomes obtained from Jurkat T cells transfected with control or LC3 siRNA. Graph: Nanoparticle concentrations in the exosomal fractions (mean, n = 2). Western blots are representative out of three independent experiments$
Figure Legend Snippet: Mitochondrial components segregate into MVBs and are secreted in exosomes. a Confocal co-localization analysis in HEK293 cells treated with FCCP (4 h) and stained with SSBP1 (red), EEA1 and CD63 (green). Bar, 10 µm. Graph shows means for Mander’s co-localization coefficient for SSBP1 and EEA1 or CD63 ( n = 3). t -test: *** P -value < 0.0001. b Confocal co-localization analysis in HEK293 cells co-transfected with Rab7-Q67L-GFP (Rab7mutGFP, green) and mitoDsRed (red), and immunostained for TFAM (purple) and nuclei (HOECHST 58, blue). Center and right images show high magnification of left images (Rab7-Q67L-GFP and mitoDsRed in upper panels; Rab7-Q67L-GFP and TFAM in lower panels). Charts: Fluorescence profiles along the corresponding white lines. Bar, 10 µm. c Mitochondrial components in the exosome fraction obtained from equal numbers of shControl and shnSMase2 J77 T cells. ATAD3, TFAM, and CD63 were detected by immunoblot; mtDNA was detected by PCR for HVRII . Graph: Quantification of exosomal proteins and mtDNA in a representative experiment ( n = 3). d Flow cytometry analysis of mitochondrial mass (Mitotracker green), intracellular ROS levels (DCFDA staining), oxidized DNA (8-OHdG Ab staining), and endogenous TFAM in HEK293 cells knocked down for nSMase2 and Rab27a. Graphs: Quantification from 5–8 independent experiments (Mean). t -test * P -value < 0.05; ** P -value < 0.001. e Electron microscopy images show defects in mitochondrial ultrastructure and cristae organization in shnSMase2 and shRab27a HEK293 cells. Graph: Quantification of mitochondrial cristae width (Mean). t -test: *** P -value < 0.0001. f Graph: Basal oxygen consumption rate (OCR) of control, shnSMase2 and shRab27a HEK293 cells. Dots represent mean from three independent experiments run in duplicate or triplicate. t -test, * P -value < 0.05, *** P -value < 0.0001. Chart: OCR from shControl, shnSMase2 and shRab27a HEK293 cells in response to oligomycin (Oligo), fccp, and rotenone plus antimycin A (Rot/AA). ( n = 2; mean ± S.E.M.). g Western blot analysis of exosomes from Jurkat T cells left untreated, serum-starved overnight or treated with bafilomycin A. Membranes were blotted for TFAM, CD81, and CD63. Graph: Nanoparticle concentration in the exosomal fractions (mean, two independent experiments). h Western blot analysis of exosomes obtained from Jurkat T cells transfected with control or LC3 siRNA. Graph: Nanoparticle concentrations in the exosomal fractions (mean, n = 2). Western blots are representative out of three independent experiments

Techniques Used: Staining, Transfection, Fluorescence, Western Blot, Flow Cytometry, Electron Microscopy, Control, Concentration Assay

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Journal: Nature Communications

Article Title: Priming of dendritic cells by DNA-containing extracellular vesicles from activated T cells through antigen-driven contacts

doi: 10.1038/s41467-018-05077-9

Figure Lengend Snippet: Mitochondrial components segregate into MVBs and are secreted in exosomes. a Confocal co-localization analysis in HEK293 cells treated with FCCP (4 h) and stained with SSBP1 (red), EEA1 and CD63 (green). Bar, 10 µm. Graph shows means for Mander’s co-localization coefficient for SSBP1 and EEA1 or CD63 ( n = 3). t -test: *** P -value < 0.0001. b Confocal co-localization analysis in HEK293 cells co-transfected with Rab7-Q67L-GFP (Rab7mutGFP, green) and mitoDsRed (red), and immunostained for TFAM (purple) and nuclei (HOECHST 58, blue). Center and right images show high magnification of left images (Rab7-Q67L-GFP and mitoDsRed in upper panels; Rab7-Q67L-GFP and TFAM in lower panels). Charts: Fluorescence profiles along the corresponding white lines. Bar, 10 µm. c Mitochondrial components in the exosome fraction obtained from equal numbers of shControl and shnSMase2 J77 T cells. ATAD3, TFAM, and CD63 were detected by immunoblot; mtDNA was detected by PCR for HVRII . Graph: Quantification of exosomal proteins and mtDNA in a representative experiment ( n = 3). d Flow cytometry analysis of mitochondrial mass (Mitotracker green), intracellular ROS levels (DCFDA staining), oxidized DNA (8-OHdG Ab staining), and endogenous TFAM in HEK293 cells knocked down for nSMase2 and Rab27a. Graphs: Quantification from 5–8 independent experiments (Mean). t -test * P -value < 0.05; ** P -value < 0.001. e Electron microscopy images show defects in mitochondrial ultrastructure and cristae organization in shnSMase2 and shRab27a HEK293 cells. Graph: Quantification of mitochondrial cristae width (Mean). t -test: *** P -value < 0.0001. f Graph: Basal oxygen consumption rate (OCR) of control, shnSMase2 and shRab27a HEK293 cells. Dots represent mean from three independent experiments run in duplicate or triplicate. t -test, * P -value < 0.05, *** P -value < 0.0001. Chart: OCR from shControl, shnSMase2 and shRab27a HEK293 cells in response to oligomycin (Oligo), fccp, and rotenone plus antimycin A (Rot/AA). ( n = 2; mean ± S.E.M.). g Western blot analysis of exosomes from Jurkat T cells left untreated, serum-starved overnight or treated with bafilomycin A. Membranes were blotted for TFAM, CD81, and CD63. Graph: Nanoparticle concentration in the exosomal fractions (mean, two independent experiments). h Western blot analysis of exosomes obtained from Jurkat T cells transfected with control or LC3 siRNA. Graph: Nanoparticle concentrations in the exosomal fractions (mean, n = 2). Western blots are representative out of three independent experiments

Article Snippet: The following plasmids were used: mitoDsRed and mitoYFP, (kindly provided by Prof. L. Scorrano, Venetian Institute for Molecular Medicine, Italy), TFAM-GFP and TFAM-DsRED (generated by inserting the human TFAM cDNA into the pEGFP and DsRED plasmids), EGFP-Rab7A Q67L, LC3-GFP and ATG5-GFP (obtained from Addgene: plasmids 28049, 24920 and 22952, respectively), shRNA pLKO control plasmid, shnSMase2 and shRab27a shRNAs (obtained from Open Biosystems), and CD63-GFP .

Techniques: Staining, Transfection, Fluorescence, Western Blot, Flow Cytometry, Electron Microscopy, Control, Concentration Assay