HRP Anti-SDHA antibody [2E3GC12FB2AE2]

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - HRP Anti-SDHA antibody [2E3GC12FB2AE2] (AB198493)

IHC image of SDHA staining in a section of formalin-fixed paraffin-embedded normal human colon*, performed on a Leica BOND. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20mins. The section was then incubated with ab198493, 1/100 dilution, for 15 mins at room temperature. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX. The inset negative control image is taken from an identical assay without primary antibody.

For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.

*Tissue obtained from the Human Research Tissue Bank, supported by the NIHR Cambridge Biomedical Research Centre.

• WB

Lab

Western blot - HRP Anti-SDHA antibody [2E3GC12FB2AE2] (AB198493)

This blot was produced using a 4-12% Bis-tris gel under the MOPS buffer system. The gel was run at 200V for 50 minutes before being transferred onto a Nitrocellulose membrane at 30V for 70 minutes. The membrane was then blocked for an hour using 3% milk before being incubated with ab198493 overnight at 4°C. Antibody binding was visualised using ECL development solution ab133406.

All lanes:

Western blot - HRP Anti-SDHA antibody [2E3GC12FB2AE2] (ab198493) at 1/5000 dilution

Lane 1:

Human heart tissue lysate - mitochondrial extract (<a href='/en-us/products/unavailable/human-heart-tissue-lysate-mitochondrial-extract-ab110337'>ab110337</a>) at 10 µg

Lane 2:

Human heart tissue lysate - mitochondrial extract (<a href='/en-us/products/unavailable/human-heart-tissue-lysate-mitochondrial-extract-ab110337'>ab110337</a>) at 5 µg

Predicted band size: 72 kDa

Observed band size: 31 kDa,42 kDa,72 kDa

true

Exposure time: 10s

• WB

Lab

Western blot - HRP Anti-SDHA antibody [2E3GC12FB2AE2] (AB198493)

ab198493 was shown to specifically react with SDHA in wild-type HEK-293 cells as signal was lost in SDHA knockout cells. Wild-type and SDHA knockout samples were subjected to SDS-PAGE. ab198493 and ab181602 (Rabbit monoclonal to GAPDH - Loading Control loading) were incubated overnight at 4°C at 1/5000 dilution and 1/20000 dilution respectively. The loading control was imaged using the Licor Odyssey CLx prior to blots being developed with ECL technique.

All lanes:

Western blot - HRP Anti-SDHA antibody [2E3GC12FB2AE2] (ab198493) at 1/5000 dilution

Lane 1:

Wild-type HEK-293 (Human epithelial cell line from embryonic kidney) whole cell lysate at 20 µg

Lane 2:

SDHA knockout HEK-293 (Human epithelial cell line from embryonic kidney) whole cell lysate at 20 µg

Lane 2:

Western blot - Human SDHA knockout HEK-293 cell line (<a href='/en-us/products/cell-lines/human-sdha-knockout-hek-293-cell-line-ab261853'>ab261853</a>)

Lane 3:

MCF7 (Human breast adenocarcinoma cell line) whole cell lysate at 20 µg

Lane 4:

Hep G2 (Human liver hepatocellular carcinoma cell line) whole cell lysate at 20 µg

Predicted band size: 72 kDa

false