Recombinant Anti-mtTFA antibody [RM1035] (ab307302)

Blocking and diluting buffer and concentration: 5% NFDM/TBST  

 Exposure time: 70 seconds

Immunohistochemical analysis of paraffin-embedded human colon tissue labeling mtTFA with AB307302 at 1/1000 (0.54 ug/ml) followed by ready to use Leica DS9800 (Bond^® Polymer Refine Detection) kit. Positive staining on mitochondria in human colon. The section was incubated with ab307302 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND^® RX instrument. Counterstained with Hematoxylin.

Secondary antibody only control: Secondary antibody is ready to use Leica DS9800 (Bond^® Polymer Refine Detection) kit.

Heat mediated antigen retrieval with Citrate buffer (pH 6.0, epitope retrieval solution 1) for 20 mins

Immunohistochemical analysis of paraffin-embedded human lung carcinoma tissue labeling mtTFA with AB307302 at 1/1000 (0.54 µg/ml) followed by ready to use Leica DS9800 (Bond^® Polymer Refine Detection) kit. Positive staining on mitochondria in human lung carcinoma. The section was incubated with ab307302 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND^® RX instrument. Counterstained with Hematoxylin.

Secondary antibody only control: Secondary antibody is ready to use Leica DS9800 (Bond^® Polymer Refine Detection) kit.

Heat mediated antigen retrieval with Citrate buffer (pH 6.0, epitope retrieval solution 1) for 20 mins

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized U-2 OS (human bone osteosarcoma epithelial cell) cells labeling mtTFA with AB307302 at 1/100 (5.4 µg/ml) dilution, followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) preadsorbed antibody at 1/1000 (2 µg/ml) dilution (Green). Confocal image showing mitochondrial staining in U-2 OS cell line. Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8). ab197491 Anti-COX IV Mouse monoclonal antibody (Alexa Fluor^® 647) - Mitochondrial Marker was used to counterstain tubulin at 1/200 dilution (2.5 µg/ml) dilution (Red). The nuclear counterstain was DAPI (Blue).

Secondary antibody only control: Secondary antibody is ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) preadsorbed at 1/1000 (2 µg/ml) dilution.

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized C2C12 (mouse myoblasts myoblast) cells labeling mtTFA with AB307302 at 1/100 (5.4 µg/ml) dilution, followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) preadsorbed antibody at 1/1000 (2 µg/ml) dilution (Green). Confocal image showing cytoplasmic staining in C2C12 cell line. Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8). ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor^® 594) was used to counterstain tubulin at 1/200 dilution (2.5 µg/ml) dilution (Red). The nuclear counterstain was DAPI (Blue).

Secondary antibody only control: Secondary antibody is ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) preadsorbed at 1/1000 (2 µg/ml) dilution.

Flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized MCF7 (human breast adenocarcinoma epithelial cell) cells labeling mtTFA with AB307302 at 1/500 dilution (0.1µg) (Red) compared with a Rabbit monoclonal IgG (ab172730) (Black) isotype control and an unlabeled control (cells without incubation with primary antibody and secondary antibody) (Blue). A Goat Anti-Rabbit IgG (Alexa Fluor^® 488, ab150081) at 1/2000 dilution was used as the secondary antibody.

Flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized C2C12 (mouse myoblast) cells labeling mtTFA with AB307302 at 1/500 dilution (0.1µg) (Red) compared with a Rabbit monoclonal IgG (ab172730) (Black) isotype control and an unlabeled control (cells without incubation with primary antibody and secondary antibody) (Blue). A Goat Anti-Rabbit IgG (Alexa Fluor^® 488, ab150081) at 1/2000 dilution was used as the secondary antibody.

mtTFA was immunoprecipitated from 0.35 mg MCF7 (human breast adenocarcinoma epithelial cell) whole cell lysate 10µg with AB307302 at 1/30 dilution (2µg in 0.35mg lysates). Western blot was performed on the immunoprecipitate using AB307302 at 1/1000 dilution. VeriBlot for IP secondary antibody (HRP) (ab131366) was used at 1/5000 dilution.

Lane 1: MCF7 (human breast adenocarcinoma epithelial cell) whole cell lysate 10µg

Lane 2: AB307302 IP in MCF7 whole cell lysate

Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab307302 in MCF7 whole cell lysate

Blocking and dilution buffer and concentration: 5% NFDM/TBST.

Exposure time: 8 seconds

mtTFA was immunoprecipitated from 0.35 mg C2C12 (mouse myoblast) whole cell lysate 10µg with AB307302 at 1/30 dilution (2µg in 0.35mg lysates). Western blot was performed on the immunoprecipitate using AB307302 at 1/1000 dilution. VeriBlot for IP secondary antibody (HRP) (ab131366) was used at 1/5000 dilution.

Lane 1: C2C12 (mouse myoblast) whole cell lysate 10µg

Lane 2: AB307302 IP in C2C12 whole cell lysate

Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab307302 in C2C12 whole cell lysate

Blocking and dilution buffer and concentration: 5% NFDM/TBST.

Exposure time: 24 seconds