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phred  (Addgene inc)


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    Addgene inc phred
    Vinculin depletion increases bioenergetics <t>of</t> <t>MDA-MB-231</t> cells. (A) Representative western blot of MDA-MB-231 cells expressing PercevalHR and <t>pHRed</t> probes transfected with siRNA to knock down vinculin (siVcl), including a scrambled control (siCtrl). (B) Quantification of the PercevalHR ratiometric signal for MDA cells expressing PercevalHR and pHRed probes treated with siRNA to knock down vinculin, including a scrambled control (siCtrl) and siVcl ( N = 3, n = 91–92 cells). (C) Representative ratiometric heatmaps of siCtrl and siVcl MDA cells expressing PercevalHR and pHRed. Scale bar, 100 µm. (D) Mean intensity of 2-NBDG of cells on 2D glass surfaces ( N = 3, n = 70–89 cells). (E) Representative images of 2-NBDG staining displayed using a fluorescent channel (top) and the Fire lookup table (bottom) to indicate the relative intensity of the fluorescent signal. Scale bar, 50 µm. (F) Mean intensity of TMRM of cells on 2D glass surfaces ( N = 3, n = 58–84 cells). (G) Representative images of TMRM staining displayed using a fluorescent channel (top) and the Fire lookup table (bottom) to indicate the relative intensity of the fluorescent signal. Scale bar, 50 µm. (H) Mean intensity of 2-NBDG for cells embedded in a 1.5 mg/ml collagen matrix ( N = 3, n = 91–103 cells). (I) Representative images of 2-NBDG staining in a 1.5 mg/ml collagen matrix displayed using a fluorescent channel (top) and the Fire lookup table (bottom) to indicate the relative intensity of the fluorescent signal. Scale bar, 20 µm. (J) Mean intensity of TMRM for cells embedded in a 1.5 mg/ml collagen matrix ( N = 3, n = 91 cells). (K) Representative images of TMRM staining in a 1.5 mg/ml collagen matrix displayed using a fluorescent channel (top) and the Fire lookup table (bottom) to indicate the relative intensity of the fluorescent signal. Scale bar, 20 µm. (L) Mean intensity of 2-NBDG in NIH/3T3 cells on 2D glass surfaces ( N = 3, n = 89–91 cells). (M) Mean intensity of TMRM in NIH/3T3 cells on 2D glass surfaces ( N = 3, n = 90–94 cells). (N) Mean intensity of 2-NBDG in MCF10A cells on 2D glass surfaces ( N = 3, n = 88–90 cells). (O) Mean intensity of TMRM in MCF10A cells on 2D glass surfaces ( N = 3, n = 90–96 cells). (P) Mean intensity of 2-NBDG for MDA ( N = 3, n = 93–98 cells) or Vcl KO ( N = 3, n = 79–86 cells) cells grown either on a tissue culture–treated surface (Adherent) or in a nonadherent plate (Floating), including representative phase-contrast images of MDA or Vcl KO cells seeded on a tissue culture surface or suspended in an ultra-low attachment plate. Scale bar, 50 µm. The box-and-whisker plot shows median and 25th/75th percentile (box), min to max (whiskers), and mean (+). Bar graphs denote the mean ± SEM. ns = not significant, **P < 0.01, ***P < 0.001, ****P < 0.0001. siVcl, siRNA targeting vinculin. Source data are available for this figure: .
    Phred, supplied by Addgene inc, used in various techniques. Bioz Stars score: 93/100, based on 14 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    phred - by Bioz Stars, 2026-04
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    Images

    1) Product Images from "Mechanometabolism of cell adhesion: Vinculin regulates bioenergetics via RhoA-ROCK"

    Article Title: Mechanometabolism of cell adhesion: Vinculin regulates bioenergetics via RhoA-ROCK

    Journal: The Journal of Cell Biology

    doi: 10.1083/jcb.202504025

    Vinculin depletion increases bioenergetics of MDA-MB-231 cells. (A) Representative western blot of MDA-MB-231 cells expressing PercevalHR and pHRed probes transfected with siRNA to knock down vinculin (siVcl), including a scrambled control (siCtrl). (B) Quantification of the PercevalHR ratiometric signal for MDA cells expressing PercevalHR and pHRed probes treated with siRNA to knock down vinculin, including a scrambled control (siCtrl) and siVcl ( N = 3, n = 91–92 cells). (C) Representative ratiometric heatmaps of siCtrl and siVcl MDA cells expressing PercevalHR and pHRed. Scale bar, 100 µm. (D) Mean intensity of 2-NBDG of cells on 2D glass surfaces ( N = 3, n = 70–89 cells). (E) Representative images of 2-NBDG staining displayed using a fluorescent channel (top) and the Fire lookup table (bottom) to indicate the relative intensity of the fluorescent signal. Scale bar, 50 µm. (F) Mean intensity of TMRM of cells on 2D glass surfaces ( N = 3, n = 58–84 cells). (G) Representative images of TMRM staining displayed using a fluorescent channel (top) and the Fire lookup table (bottom) to indicate the relative intensity of the fluorescent signal. Scale bar, 50 µm. (H) Mean intensity of 2-NBDG for cells embedded in a 1.5 mg/ml collagen matrix ( N = 3, n = 91–103 cells). (I) Representative images of 2-NBDG staining in a 1.5 mg/ml collagen matrix displayed using a fluorescent channel (top) and the Fire lookup table (bottom) to indicate the relative intensity of the fluorescent signal. Scale bar, 20 µm. (J) Mean intensity of TMRM for cells embedded in a 1.5 mg/ml collagen matrix ( N = 3, n = 91 cells). (K) Representative images of TMRM staining in a 1.5 mg/ml collagen matrix displayed using a fluorescent channel (top) and the Fire lookup table (bottom) to indicate the relative intensity of the fluorescent signal. Scale bar, 20 µm. (L) Mean intensity of 2-NBDG in NIH/3T3 cells on 2D glass surfaces ( N = 3, n = 89–91 cells). (M) Mean intensity of TMRM in NIH/3T3 cells on 2D glass surfaces ( N = 3, n = 90–94 cells). (N) Mean intensity of 2-NBDG in MCF10A cells on 2D glass surfaces ( N = 3, n = 88–90 cells). (O) Mean intensity of TMRM in MCF10A cells on 2D glass surfaces ( N = 3, n = 90–96 cells). (P) Mean intensity of 2-NBDG for MDA ( N = 3, n = 93–98 cells) or Vcl KO ( N = 3, n = 79–86 cells) cells grown either on a tissue culture–treated surface (Adherent) or in a nonadherent plate (Floating), including representative phase-contrast images of MDA or Vcl KO cells seeded on a tissue culture surface or suspended in an ultra-low attachment plate. Scale bar, 50 µm. The box-and-whisker plot shows median and 25th/75th percentile (box), min to max (whiskers), and mean (+). Bar graphs denote the mean ± SEM. ns = not significant, **P < 0.01, ***P < 0.001, ****P < 0.0001. siVcl, siRNA targeting vinculin. Source data are available for this figure: .
    Figure Legend Snippet: Vinculin depletion increases bioenergetics of MDA-MB-231 cells. (A) Representative western blot of MDA-MB-231 cells expressing PercevalHR and pHRed probes transfected with siRNA to knock down vinculin (siVcl), including a scrambled control (siCtrl). (B) Quantification of the PercevalHR ratiometric signal for MDA cells expressing PercevalHR and pHRed probes treated with siRNA to knock down vinculin, including a scrambled control (siCtrl) and siVcl ( N = 3, n = 91–92 cells). (C) Representative ratiometric heatmaps of siCtrl and siVcl MDA cells expressing PercevalHR and pHRed. Scale bar, 100 µm. (D) Mean intensity of 2-NBDG of cells on 2D glass surfaces ( N = 3, n = 70–89 cells). (E) Representative images of 2-NBDG staining displayed using a fluorescent channel (top) and the Fire lookup table (bottom) to indicate the relative intensity of the fluorescent signal. Scale bar, 50 µm. (F) Mean intensity of TMRM of cells on 2D glass surfaces ( N = 3, n = 58–84 cells). (G) Representative images of TMRM staining displayed using a fluorescent channel (top) and the Fire lookup table (bottom) to indicate the relative intensity of the fluorescent signal. Scale bar, 50 µm. (H) Mean intensity of 2-NBDG for cells embedded in a 1.5 mg/ml collagen matrix ( N = 3, n = 91–103 cells). (I) Representative images of 2-NBDG staining in a 1.5 mg/ml collagen matrix displayed using a fluorescent channel (top) and the Fire lookup table (bottom) to indicate the relative intensity of the fluorescent signal. Scale bar, 20 µm. (J) Mean intensity of TMRM for cells embedded in a 1.5 mg/ml collagen matrix ( N = 3, n = 91 cells). (K) Representative images of TMRM staining in a 1.5 mg/ml collagen matrix displayed using a fluorescent channel (top) and the Fire lookup table (bottom) to indicate the relative intensity of the fluorescent signal. Scale bar, 20 µm. (L) Mean intensity of 2-NBDG in NIH/3T3 cells on 2D glass surfaces ( N = 3, n = 89–91 cells). (M) Mean intensity of TMRM in NIH/3T3 cells on 2D glass surfaces ( N = 3, n = 90–94 cells). (N) Mean intensity of 2-NBDG in MCF10A cells on 2D glass surfaces ( N = 3, n = 88–90 cells). (O) Mean intensity of TMRM in MCF10A cells on 2D glass surfaces ( N = 3, n = 90–96 cells). (P) Mean intensity of 2-NBDG for MDA ( N = 3, n = 93–98 cells) or Vcl KO ( N = 3, n = 79–86 cells) cells grown either on a tissue culture–treated surface (Adherent) or in a nonadherent plate (Floating), including representative phase-contrast images of MDA or Vcl KO cells seeded on a tissue culture surface or suspended in an ultra-low attachment plate. Scale bar, 50 µm. The box-and-whisker plot shows median and 25th/75th percentile (box), min to max (whiskers), and mean (+). Bar graphs denote the mean ± SEM. ns = not significant, **P < 0.01, ***P < 0.001, ****P < 0.0001. siVcl, siRNA targeting vinculin. Source data are available for this figure: .

    Techniques Used: Western Blot, Expressing, Transfection, Knockdown, Control, Staining, Whisker Assay

    Representative western blots. (A) Representative western blot confirming Vcl knockdown in MDA-MB-231 cells expressing PercevalHR and pHRed probes, including a scrambled control (siCtrl) and siVcl. Relevant samples are outlined with a red box, and uncropped membrane is included to show the ladder. (B and C) Representative western blot and (C) quantification from three independent western blots confirming Vcl KO and rescue in MDA-MB-231 cells ( N = 3). The Pre-Sort population refers to the Vcl KO cells transduced with Vcl-Venus before positively expressing Venus cells were sorted using FACS. Bar graph denotes the mean ± SEM. ns = not significant. (D) Representative western blot to confirm Vcl KO in NIH/3T3 cells. (E) Representative western blot to confirm Vcl KO in MCF10A cells. siVcl, siRNA targeting vinculin; FACS, fluorescence-activated cell sorting. Source data are available for this figure: .
    Figure Legend Snippet: Representative western blots. (A) Representative western blot confirming Vcl knockdown in MDA-MB-231 cells expressing PercevalHR and pHRed probes, including a scrambled control (siCtrl) and siVcl. Relevant samples are outlined with a red box, and uncropped membrane is included to show the ladder. (B and C) Representative western blot and (C) quantification from three independent western blots confirming Vcl KO and rescue in MDA-MB-231 cells ( N = 3). The Pre-Sort population refers to the Vcl KO cells transduced with Vcl-Venus before positively expressing Venus cells were sorted using FACS. Bar graph denotes the mean ± SEM. ns = not significant. (D) Representative western blot to confirm Vcl KO in NIH/3T3 cells. (E) Representative western blot to confirm Vcl KO in MCF10A cells. siVcl, siRNA targeting vinculin; FACS, fluorescence-activated cell sorting. Source data are available for this figure: .

    Techniques Used: Western Blot, Knockdown, Expressing, Control, Membrane, Transduction, Fluorescence, FACS



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    Vinculin depletion increases bioenergetics <t>of</t> <t>MDA-MB-231</t> cells. (A) Representative western blot of MDA-MB-231 cells expressing PercevalHR and <t>pHRed</t> probes transfected with siRNA to knock down vinculin (siVcl), including a scrambled control (siCtrl). (B) Quantification of the PercevalHR ratiometric signal for MDA cells expressing PercevalHR and pHRed probes treated with siRNA to knock down vinculin, including a scrambled control (siCtrl) and siVcl ( N = 3, n = 91–92 cells). (C) Representative ratiometric heatmaps of siCtrl and siVcl MDA cells expressing PercevalHR and pHRed. Scale bar, 100 µm. (D) Mean intensity of 2-NBDG of cells on 2D glass surfaces ( N = 3, n = 70–89 cells). (E) Representative images of 2-NBDG staining displayed using a fluorescent channel (top) and the Fire lookup table (bottom) to indicate the relative intensity of the fluorescent signal. Scale bar, 50 µm. (F) Mean intensity of TMRM of cells on 2D glass surfaces ( N = 3, n = 58–84 cells). (G) Representative images of TMRM staining displayed using a fluorescent channel (top) and the Fire lookup table (bottom) to indicate the relative intensity of the fluorescent signal. Scale bar, 50 µm. (H) Mean intensity of 2-NBDG for cells embedded in a 1.5 mg/ml collagen matrix ( N = 3, n = 91–103 cells). (I) Representative images of 2-NBDG staining in a 1.5 mg/ml collagen matrix displayed using a fluorescent channel (top) and the Fire lookup table (bottom) to indicate the relative intensity of the fluorescent signal. Scale bar, 20 µm. (J) Mean intensity of TMRM for cells embedded in a 1.5 mg/ml collagen matrix ( N = 3, n = 91 cells). (K) Representative images of TMRM staining in a 1.5 mg/ml collagen matrix displayed using a fluorescent channel (top) and the Fire lookup table (bottom) to indicate the relative intensity of the fluorescent signal. Scale bar, 20 µm. (L) Mean intensity of 2-NBDG in NIH/3T3 cells on 2D glass surfaces ( N = 3, n = 89–91 cells). (M) Mean intensity of TMRM in NIH/3T3 cells on 2D glass surfaces ( N = 3, n = 90–94 cells). (N) Mean intensity of 2-NBDG in MCF10A cells on 2D glass surfaces ( N = 3, n = 88–90 cells). (O) Mean intensity of TMRM in MCF10A cells on 2D glass surfaces ( N = 3, n = 90–96 cells). (P) Mean intensity of 2-NBDG for MDA ( N = 3, n = 93–98 cells) or Vcl KO ( N = 3, n = 79–86 cells) cells grown either on a tissue culture–treated surface (Adherent) or in a nonadherent plate (Floating), including representative phase-contrast images of MDA or Vcl KO cells seeded on a tissue culture surface or suspended in an ultra-low attachment plate. Scale bar, 50 µm. The box-and-whisker plot shows median and 25th/75th percentile (box), min to max (whiskers), and mean (+). Bar graphs denote the mean ± SEM. ns = not significant, **P < 0.01, ***P < 0.001, ****P < 0.0001. siVcl, siRNA targeting vinculin. Source data are available for this figure: .
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    phred-like quality score threshold  (10X Genomics)
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    10X Genomics phred-like quality score threshold
    Vinculin depletion increases bioenergetics <t>of</t> <t>MDA-MB-231</t> cells. (A) Representative western blot of MDA-MB-231 cells expressing PercevalHR and <t>pHRed</t> probes transfected with siRNA to knock down vinculin (siVcl), including a scrambled control (siCtrl). (B) Quantification of the PercevalHR ratiometric signal for MDA cells expressing PercevalHR and pHRed probes treated with siRNA to knock down vinculin, including a scrambled control (siCtrl) and siVcl ( N = 3, n = 91–92 cells). (C) Representative ratiometric heatmaps of siCtrl and siVcl MDA cells expressing PercevalHR and pHRed. Scale bar, 100 µm. (D) Mean intensity of 2-NBDG of cells on 2D glass surfaces ( N = 3, n = 70–89 cells). (E) Representative images of 2-NBDG staining displayed using a fluorescent channel (top) and the Fire lookup table (bottom) to indicate the relative intensity of the fluorescent signal. Scale bar, 50 µm. (F) Mean intensity of TMRM of cells on 2D glass surfaces ( N = 3, n = 58–84 cells). (G) Representative images of TMRM staining displayed using a fluorescent channel (top) and the Fire lookup table (bottom) to indicate the relative intensity of the fluorescent signal. Scale bar, 50 µm. (H) Mean intensity of 2-NBDG for cells embedded in a 1.5 mg/ml collagen matrix ( N = 3, n = 91–103 cells). (I) Representative images of 2-NBDG staining in a 1.5 mg/ml collagen matrix displayed using a fluorescent channel (top) and the Fire lookup table (bottom) to indicate the relative intensity of the fluorescent signal. Scale bar, 20 µm. (J) Mean intensity of TMRM for cells embedded in a 1.5 mg/ml collagen matrix ( N = 3, n = 91 cells). (K) Representative images of TMRM staining in a 1.5 mg/ml collagen matrix displayed using a fluorescent channel (top) and the Fire lookup table (bottom) to indicate the relative intensity of the fluorescent signal. Scale bar, 20 µm. (L) Mean intensity of 2-NBDG in NIH/3T3 cells on 2D glass surfaces ( N = 3, n = 89–91 cells). (M) Mean intensity of TMRM in NIH/3T3 cells on 2D glass surfaces ( N = 3, n = 90–94 cells). (N) Mean intensity of 2-NBDG in MCF10A cells on 2D glass surfaces ( N = 3, n = 88–90 cells). (O) Mean intensity of TMRM in MCF10A cells on 2D glass surfaces ( N = 3, n = 90–96 cells). (P) Mean intensity of 2-NBDG for MDA ( N = 3, n = 93–98 cells) or Vcl KO ( N = 3, n = 79–86 cells) cells grown either on a tissue culture–treated surface (Adherent) or in a nonadherent plate (Floating), including representative phase-contrast images of MDA or Vcl KO cells seeded on a tissue culture surface or suspended in an ultra-low attachment plate. Scale bar, 50 µm. The box-and-whisker plot shows median and 25th/75th percentile (box), min to max (whiskers), and mean (+). Bar graphs denote the mean ± SEM. ns = not significant, **P < 0.01, ***P < 0.001, ****P < 0.0001. siVcl, siRNA targeting vinculin. Source data are available for this figure: .
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    Image Search Results


    Vinculin depletion increases bioenergetics of MDA-MB-231 cells. (A) Representative western blot of MDA-MB-231 cells expressing PercevalHR and pHRed probes transfected with siRNA to knock down vinculin (siVcl), including a scrambled control (siCtrl). (B) Quantification of the PercevalHR ratiometric signal for MDA cells expressing PercevalHR and pHRed probes treated with siRNA to knock down vinculin, including a scrambled control (siCtrl) and siVcl ( N = 3, n = 91–92 cells). (C) Representative ratiometric heatmaps of siCtrl and siVcl MDA cells expressing PercevalHR and pHRed. Scale bar, 100 µm. (D) Mean intensity of 2-NBDG of cells on 2D glass surfaces ( N = 3, n = 70–89 cells). (E) Representative images of 2-NBDG staining displayed using a fluorescent channel (top) and the Fire lookup table (bottom) to indicate the relative intensity of the fluorescent signal. Scale bar, 50 µm. (F) Mean intensity of TMRM of cells on 2D glass surfaces ( N = 3, n = 58–84 cells). (G) Representative images of TMRM staining displayed using a fluorescent channel (top) and the Fire lookup table (bottom) to indicate the relative intensity of the fluorescent signal. Scale bar, 50 µm. (H) Mean intensity of 2-NBDG for cells embedded in a 1.5 mg/ml collagen matrix ( N = 3, n = 91–103 cells). (I) Representative images of 2-NBDG staining in a 1.5 mg/ml collagen matrix displayed using a fluorescent channel (top) and the Fire lookup table (bottom) to indicate the relative intensity of the fluorescent signal. Scale bar, 20 µm. (J) Mean intensity of TMRM for cells embedded in a 1.5 mg/ml collagen matrix ( N = 3, n = 91 cells). (K) Representative images of TMRM staining in a 1.5 mg/ml collagen matrix displayed using a fluorescent channel (top) and the Fire lookup table (bottom) to indicate the relative intensity of the fluorescent signal. Scale bar, 20 µm. (L) Mean intensity of 2-NBDG in NIH/3T3 cells on 2D glass surfaces ( N = 3, n = 89–91 cells). (M) Mean intensity of TMRM in NIH/3T3 cells on 2D glass surfaces ( N = 3, n = 90–94 cells). (N) Mean intensity of 2-NBDG in MCF10A cells on 2D glass surfaces ( N = 3, n = 88–90 cells). (O) Mean intensity of TMRM in MCF10A cells on 2D glass surfaces ( N = 3, n = 90–96 cells). (P) Mean intensity of 2-NBDG for MDA ( N = 3, n = 93–98 cells) or Vcl KO ( N = 3, n = 79–86 cells) cells grown either on a tissue culture–treated surface (Adherent) or in a nonadherent plate (Floating), including representative phase-contrast images of MDA or Vcl KO cells seeded on a tissue culture surface or suspended in an ultra-low attachment plate. Scale bar, 50 µm. The box-and-whisker plot shows median and 25th/75th percentile (box), min to max (whiskers), and mean (+). Bar graphs denote the mean ± SEM. ns = not significant, **P < 0.01, ***P < 0.001, ****P < 0.0001. siVcl, siRNA targeting vinculin. Source data are available for this figure: .

    Journal: The Journal of Cell Biology

    Article Title: Mechanometabolism of cell adhesion: Vinculin regulates bioenergetics via RhoA-ROCK

    doi: 10.1083/jcb.202504025

    Figure Lengend Snippet: Vinculin depletion increases bioenergetics of MDA-MB-231 cells. (A) Representative western blot of MDA-MB-231 cells expressing PercevalHR and pHRed probes transfected with siRNA to knock down vinculin (siVcl), including a scrambled control (siCtrl). (B) Quantification of the PercevalHR ratiometric signal for MDA cells expressing PercevalHR and pHRed probes treated with siRNA to knock down vinculin, including a scrambled control (siCtrl) and siVcl ( N = 3, n = 91–92 cells). (C) Representative ratiometric heatmaps of siCtrl and siVcl MDA cells expressing PercevalHR and pHRed. Scale bar, 100 µm. (D) Mean intensity of 2-NBDG of cells on 2D glass surfaces ( N = 3, n = 70–89 cells). (E) Representative images of 2-NBDG staining displayed using a fluorescent channel (top) and the Fire lookup table (bottom) to indicate the relative intensity of the fluorescent signal. Scale bar, 50 µm. (F) Mean intensity of TMRM of cells on 2D glass surfaces ( N = 3, n = 58–84 cells). (G) Representative images of TMRM staining displayed using a fluorescent channel (top) and the Fire lookup table (bottom) to indicate the relative intensity of the fluorescent signal. Scale bar, 50 µm. (H) Mean intensity of 2-NBDG for cells embedded in a 1.5 mg/ml collagen matrix ( N = 3, n = 91–103 cells). (I) Representative images of 2-NBDG staining in a 1.5 mg/ml collagen matrix displayed using a fluorescent channel (top) and the Fire lookup table (bottom) to indicate the relative intensity of the fluorescent signal. Scale bar, 20 µm. (J) Mean intensity of TMRM for cells embedded in a 1.5 mg/ml collagen matrix ( N = 3, n = 91 cells). (K) Representative images of TMRM staining in a 1.5 mg/ml collagen matrix displayed using a fluorescent channel (top) and the Fire lookup table (bottom) to indicate the relative intensity of the fluorescent signal. Scale bar, 20 µm. (L) Mean intensity of 2-NBDG in NIH/3T3 cells on 2D glass surfaces ( N = 3, n = 89–91 cells). (M) Mean intensity of TMRM in NIH/3T3 cells on 2D glass surfaces ( N = 3, n = 90–94 cells). (N) Mean intensity of 2-NBDG in MCF10A cells on 2D glass surfaces ( N = 3, n = 88–90 cells). (O) Mean intensity of TMRM in MCF10A cells on 2D glass surfaces ( N = 3, n = 90–96 cells). (P) Mean intensity of 2-NBDG for MDA ( N = 3, n = 93–98 cells) or Vcl KO ( N = 3, n = 79–86 cells) cells grown either on a tissue culture–treated surface (Adherent) or in a nonadherent plate (Floating), including representative phase-contrast images of MDA or Vcl KO cells seeded on a tissue culture surface or suspended in an ultra-low attachment plate. Scale bar, 50 µm. The box-and-whisker plot shows median and 25th/75th percentile (box), min to max (whiskers), and mean (+). Bar graphs denote the mean ± SEM. ns = not significant, **P < 0.01, ***P < 0.001, ****P < 0.0001. siVcl, siRNA targeting vinculin. Source data are available for this figure: .

    Article Snippet: MDA-MB-231 cells stably co-expressing PercevalHR (#49083; Addgene) and pHRed (#31473; Addgene) probes, gifts from Gary Yellen (Harvard Medical School, Boston, MA, USA), were transfected with 50 pmol/μl nontargeting scrambled control siRNA (AM4611; Thermo Fisher Scientific) or siRNA targeting Vcl (AM51331 [siRNA ID: 108455] and AM16708 [siRNA ID: 108453]; Thermo Fisher Scientific) using Lipofectamine 2000 (11668030; Thermo Fisher Scientific) and Opti-MEM transfection medium (31985062; Thermo Fisher Scientific) according to the manufacturer’s instructions.

    Techniques: Western Blot, Expressing, Transfection, Knockdown, Control, Staining, Whisker Assay

    Representative western blots. (A) Representative western blot confirming Vcl knockdown in MDA-MB-231 cells expressing PercevalHR and pHRed probes, including a scrambled control (siCtrl) and siVcl. Relevant samples are outlined with a red box, and uncropped membrane is included to show the ladder. (B and C) Representative western blot and (C) quantification from three independent western blots confirming Vcl KO and rescue in MDA-MB-231 cells ( N = 3). The Pre-Sort population refers to the Vcl KO cells transduced with Vcl-Venus before positively expressing Venus cells were sorted using FACS. Bar graph denotes the mean ± SEM. ns = not significant. (D) Representative western blot to confirm Vcl KO in NIH/3T3 cells. (E) Representative western blot to confirm Vcl KO in MCF10A cells. siVcl, siRNA targeting vinculin; FACS, fluorescence-activated cell sorting. Source data are available for this figure: .

    Journal: The Journal of Cell Biology

    Article Title: Mechanometabolism of cell adhesion: Vinculin regulates bioenergetics via RhoA-ROCK

    doi: 10.1083/jcb.202504025

    Figure Lengend Snippet: Representative western blots. (A) Representative western blot confirming Vcl knockdown in MDA-MB-231 cells expressing PercevalHR and pHRed probes, including a scrambled control (siCtrl) and siVcl. Relevant samples are outlined with a red box, and uncropped membrane is included to show the ladder. (B and C) Representative western blot and (C) quantification from three independent western blots confirming Vcl KO and rescue in MDA-MB-231 cells ( N = 3). The Pre-Sort population refers to the Vcl KO cells transduced with Vcl-Venus before positively expressing Venus cells were sorted using FACS. Bar graph denotes the mean ± SEM. ns = not significant. (D) Representative western blot to confirm Vcl KO in NIH/3T3 cells. (E) Representative western blot to confirm Vcl KO in MCF10A cells. siVcl, siRNA targeting vinculin; FACS, fluorescence-activated cell sorting. Source data are available for this figure: .

    Article Snippet: MDA-MB-231 cells stably co-expressing PercevalHR (#49083; Addgene) and pHRed (#31473; Addgene) probes, gifts from Gary Yellen (Harvard Medical School, Boston, MA, USA), were transfected with 50 pmol/μl nontargeting scrambled control siRNA (AM4611; Thermo Fisher Scientific) or siRNA targeting Vcl (AM51331 [siRNA ID: 108455] and AM16708 [siRNA ID: 108453]; Thermo Fisher Scientific) using Lipofectamine 2000 (11668030; Thermo Fisher Scientific) and Opti-MEM transfection medium (31985062; Thermo Fisher Scientific) according to the manufacturer’s instructions.

    Techniques: Western Blot, Knockdown, Expressing, Control, Membrane, Transduction, Fluorescence, FACS