Anti-COX5B antibody [16H12H9]

• Flow Cyt

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Flow Cytometry - Anti-COX5B antibody [16H12H9] (AB110263)

Overlay histogram showing HepG2 cells stained with ab110263 (red line). The cells were fixed with 80% methanol (5 min) and then permeabilized with 0.1% PBS-Tween for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab110263, 1μg/1x10⁶ cells) for 30 min at 22°C. The secondary antibody used was DyLight® 488 goat anti-mouse IgG (H+L) (ab96879) at 1/500 dilution for 30 min at 22°C. Isotype control antibody (black line) was mouse IgG2b [PLPV219] (ab91366, 2μg/1x10⁶ cells) used under the same conditions. Acquisition of >5,000 events was performed. This antibody gave a positive signal in HepG2 cells fixed with 4% paraformaldehyde (10 min)/permeabilized with 0.1% PBS-Tween for 20 min used under the same conditions.

• WB

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Western blot - Anti-COX5B antibody [16H12H9] (AB110263)

All lanes:

Western blot - Anti-COX5B antibody [16H12H9] (ab110263) at 1 µg/mL

Lane 1:

Human heart mitochondria at 5 µg

Lane 2:

Bovine heart mitochondria at 1 µg

Lane 3:

Rat heart mitochondria at 10 µg

Lane 4:

Mouse heart mitochondria at 10 µg

Predicted band size: 14 kDa

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• WB

CiteAb

Western blot - Anti-COX5B antibody [16H12H9] (AB110263)

COX5B western blot using anti-COX5B antibody [16H12H9] ab110263. Publication image and figure legend from Das, S., Bedja, D., et al., 2014, PLoS One, PubMed 24810628.

ab110263 was used in this publication in western blot. This may not be the same as the application(s) guaranteed by Abcam. For a full list of applications guaranteed by Abcam for ab110263 please see the product overview.

Mitochondrial Complex IV Remodeling after miR-181c Treatment.(A) qPCR data show that overexpression of miR-181c significantly reduces the mRNA levels of all mitochondrial complex IV genes with 3 weeks treatment. The treatment protocol has no effect on other mitochondrial genes, such as ND2 (complex I) and ATPase 8 (complex V). Content of mRNA was first normalized to 12S rRNA, a mitochondrial gene, as 12S rRNA expression did not change with miR-181c overexpresssion. Then we normalized the data to the sham group. *p<0.05 vs. sham (n = 6). (B) Western blot shows that miR-181c overexpression significantly reduces the protein content of both mt-COX1 and mt-COX2. VDAC was used as a loading control. The data were normalized to the sham group. *p<0.05 vs. sham (n = 6). (C) Western blot shows that miR-181c overexpression has no effect on Transcription Factor A, Mitochondria (TFAM), either in the total heart homogenate (left panel) or the heart-derived mitochondrial fraction (right panel). TFAM plays an important role in mitochondrial gene transcription, by activating 3 different promoter regions in the D-loop area of the mitochondrial genome. TFAM also translocates from the cytosol to the mitochondria as part of the mitochondrial gene transcription process. α-tubulin (for total heart homogenate) and VDAC (mitochondrial fraction) were used as loading controls. The data were normalized to the sham group (n = 3). (D) Western blot shows the changes of other isoforms of mitochondrial respiratory chain complex IV. We have observed a significant decrease in the protein content of COX VIIa in the miR-181c overexpression groups, but no effect on COX 5A and COX 5B. α-tubulin was used as the loading control. The data were normalized to the sham group (n = 3).

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