Anti-Cytochrome C antibody [37BA11]

• WB

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Western blot - Anti-Cytochrome C antibody [37BA11] (AB110325)

All lanes:

Western blot - Anti-Cytochrome C antibody [37BA11] (ab110325) at 1 µg/mL

Lane 1:

Isolated mitochondria from Human heart at 5 µg

Lane 2:

Isolated mitochondria from Bovine heart at 1 µg

Lane 3:

Isolated mitochondria from Rat heart at 10 µg

Lane 4:

Isolated mitochondria from Mouse heart at 10 µg

Predicted band size: 11 kDa

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• ICC/IF

AbReview47048****

Immunocytochemistry/ Immunofluorescence - Anti-Cytochrome C antibody [37BA11] (AB110325)

ab110325 staining Cytochrome C in MDA MB 231 cells by ICC/IF (Immunocytochemistry/immunofluorescence). Cells were fixed with formaldehyde, permeabilized with 1% Triton X-100 and blocked with 10% BSA for 1 hour at 21°C. Samples were incubated with primary antibody (1/100) for 1 hour. An undiluted DyLight^® 550-conjugated goat anti-mouse IgG polyclonal was used as the secondary antibody.

This image is courtesy of an anonymous Abreview

• Flow Cyt

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Flow Cytometry - Anti-Cytochrome C antibody [37BA11] (AB110325)

Flow cytometric analysis using ab110325 at 1µg/ml staining Cytochrome C in HeLa cells (blue). Isotype control antibody (red).

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-Cytochrome C antibody [37BA11] (AB110325)

Immunocytochemistry analysis using ab110325 at 2μg/ml staining Cytochrome C in Human fibroblasts (fixed and permeabilized), followed by a fluorescent goat-anti-mouse IgG.

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-Cytochrome C antibody [37BA11] (AB110325)

ab110325 staining cytochrome C in MCF7 cells treated with 15-Deoxy-delta12,14-prostaglandin J2 (ab141717), by ICC/IF. Expression of cytochrome C changes from mitochondrial puncta to a difuse staining pattern with increased concentration of 15-Deoxy-delta12,14-prostaglandin J2, as described in literature.
The cells were incubated at 37°C for 24 hours in media containing different concentrations of ab141717 (15-Deoxy-delta12,14-prostaglandin J2) in DMSO, fixed with 100% methanol for 5 minutes at -20°C and blocked with PBS containing 10% goat serum, 0.3 M glycine, 1% BSA and 0.1% tween for 2h at room temperature. Staining of the treated cells with ab110325 (10 μg/ml) was performed overnight at 4°C in PBS containing 1% BSA and 0.1% tween. A DyLight 488 anti-mouse polyclonal antibody (ab96879) at 1/250 dilution was used as the secondary antibody. Nuclei (blue) were counterstained with DAPI and membrane is was stained using WGA (red).

• WB

CiteAb

Western blot - Anti-Cytochrome C antibody [37BA11] (AB110325)

Western Blotting using Anti-Cytochrome C antibody [37BA11], ab110325. Publication image from Yu, B. et al., 2020, Nat Commun, 32439975. Legend direct from paper.

Mitochondrial dynamics and function change upon PGAM5 deletion.a Mitochondrial morphology outlined by Tom20 antibodies in control and PGAM5−/− ARPE-19 cells. Scale bar = 20 µm. n = 5. b Western blots showing upregulation of phosphor-Drp1(S637) but not total Drp1 in PGAM5−/− ARPE-19 cells.α-Tubulin was used as loading control. n = 4. c Western blots showing PGAM5 cleavage and Drp1(S637) dephosphorylation in ARPE-19 cells by CCCP treatment. n = 3. d Co-immunoprecipitation experiment using Axin1 antibody, showing that Axin1 interacts with both Drp1 and cleaved PGAM5 in ARPE-19 cells. n = 3. e Western blots showing upregulation of mitochondrial proteins (Tom20, CYTC, CYPD) and downregulation of PGC1α in PGAM5−/− ARPE-19 cells.α-Tubulin was used as loading control. n = 4. f Increased mitochondrial DNA in PGAM5−/− ARPE-19 cells. n = 5. *p = 0.0111, two-tailed unpaired t-tests; error bars, mean ± s.e.m. g Increased mitochondrial protein Cypd in the RPE/choroid of Pgam5−/− mice.α-Tubulin was used as loading control. n = 3. h Decreased mitochondrial turnover in PGAM5−/− ARPE-19 by MitoTimer transfection and labeling. Scale Bar equals to 20 µm. n = 3. i Mitochondrial membrane potential change as labeled by JC-1 in WT and PGAM5−/− ARPE-19 cells. Scale bar = 20 µm. n = 5. j ATP level change as measured in short-term (1 week) and long-term (8 weeks) culture of WT and PGAM5−/− ARPE-19 cells. n = 3, ****p < 0.0001, two-way ANOVA Tukey’s multiple comparisons test; error bars, mean ± s.e.m. k ROS change as measured in short-term (1 week) and long-term (8 weeks) culture of WT and PGAM5−/− ARPE-19 cells. n = 3, *p < 0.05; ****p < 0.0001, two-way ANOVA Tukey’s multiple comparisons test. n.s. represents no significance. Error bars, mean ± s.e.m.; for assays in the figure, n represents the number of biologically independent experiments. Images were captured under same settings, and representative images were shown. Source data are available as a Source Data file.

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• WB

CiteAb

Western blot - Anti-Cytochrome C antibody [37BA11] (AB110325)

Western Blotting using Anti-Cytochrome C antibody [37BA11], ab110325. Publication image from Yu, B. et al., 2020, Nat Commun, 32439975. Legend direct from paper.

Rescue of PGAM5−/− accelerated senescence phenotype by Drp-S637A overexpression.a Mitochondrial morphology outlined by Tom20 antibodies in WT and PGAM5−/− ARPE-19 cells overexpressing GFP or DRP1 S637A, respectively. Scale bar = 20 µm. n = 5. b Quantification of mitochondrial branch in a. n = 5, ***p = 0.0007, ****p < 0.0001, n.s. no significant. two-way ANOVA Tukey’s multiple comparisons test. Error bars, mean ± s.d. c Upregulation of mitochondrial proteins (Tom20, CYTC, CYPD) were rescued by Drp1 S637A overexpression in PGAM5−/− ARPE-19 cells.α-Tubulin was used as loading control. n = 4. d Expression of phosphor-AMPK, phosphor-TSC2, total-TSC2, phosphor-S6, total S6 and Drp1 in WT and PGAM5−/− ARPE-19 cells transduced with GFP or Drp1 S637A expressing lentivirus. β-Actin was used as loading control. n = 4. e ATP level in short-term (1 week) culture of WT and PGAM5−/− ARPE-19 cells transduced with GFP or Drp1 S637A expressing lentivirus. n = 9 biologically independent samples. ***p = 0.0005, ****p < 0.0001, two-way ANOVA Tukey’s multiple comparisons test. Error bars, mean ± s.d. f ATP level in long-term (8 week) culture of WT and PGAM5−/− ARPE-19 cells transduced with GFP or Drp1 S637A expressing lentivirus. n = 9 biologically independent samples. ****p < 0.0001, two-way ANOVA Tukey’s multiple comparisons test. Error bars, mean ± s.d. g Partial rescue of senescence phenotype in PGAM5−/− ARPE-19 cells by Drp1 S637A overexpression as shown by SA-β-Gal assay. Scale bar = 100 µm. n = 4. h Quantification β-gal-positive cells in g. n = 4. ****p < 0.0001, two-way ANOVA Tukey’s multiple comparisons test. Error bars, mean ± s.d. For assays in the figure, n represents the number of biologically independent experiments unless otherwise specified. Images were captured under same settings, and representative images were shown. Source data are available as a Source Data file.

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• WB

CiteAb

Western blot - Anti-Cytochrome C antibody [37BA11] (AB110325)

Western Blotting using Anti-Cytochrome C antibody [37BA11], ab110325. Publication image from Torralba, D. et al., 2018, Nat Commun, 29985392. Legend direct from paper.

Exosomes shuttle mitochondrial proteins. a Differential ultracentrifugation protocol for isolating and purifying exosomes from cell culture supernatants. b Size distribution analysis by Nanoparticle Tracking Analysis (NTA) of purified EVs from primary human T lymphoblasts. c Gene ontology (GO) cellular component analysis of peptides identified in T cell EVs. d Western blot analysis of mitochondrial proteins in EVs obtained from the culture supernatant of primary human T lymphoblasts with or without treatment with phorbol myristate acetate (PMA) plus ionomycin. Cells and EVs were blotted for proteins associated with mtDNA (TFAM and ATAD3), for proteins located in the inner mitochondrial membrane (COX1 and Cytochrome C), the outer mitochondrial membrane (VDAC1), the mitochondrial matrix (mitochondrial manganese superoxide dismutase, MnSOD), and for the exosome markers TSG101 and CD81. e Western blot analysis of ATAD3, TFAM, and the exosome markers CD81 and TSG101 in sucrose fractions. EVs obtained from the culture supernatant of human T lymphoblasts were laid on a discontinuous sucrose gradient and floated by overnight centrifugation. Gradient fractions were collected and analyzed by immunoblot to reveal the distribution of mtDNA-binding proteins and exosomal proteins in the sucrose fractions from lower to higher sucrose density (left to right). Gels shown are representative out of three independent experiments

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