Anti-Lipoamide Dehydrogenase antibody [EPR6635]

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Lipoamide Dehydrogenase antibody [EPR6635] (AB133551)

Immunohistochemical analysis of paraffin embedded Human kidney tissue labelling Lipoamide Dehydrogenase with ab133551 antibody at a dilution of 1/100.

Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Lipoamide Dehydrogenase antibody [EPR6635] (AB133551)

Immunohistochemical analysis of paraffin embedded Human colon tissue labelling Lipoamide Dehydrogenase with ab133551 antibody at a dilution of 1/100.

Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-Lipoamide Dehydrogenase antibody [EPR6635] (AB133551)

Immunocytochemistry/ Immunofluorescence analysis of Jurkat (human T cell leukemia T lymphocyte from peripheral blood) labeling Lipoamide Dehydrogenase with ab133551 at 1/200 dilution followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed secondary antibody at 1/1000 dilution. Cells were fixed with 4% Paraformaldehyde and permeabilised with 0.1% TritonX-100. ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor® 594) at 1/200 was used as the counterstain antibody. Nuclear counter stain was DAPI. Confocal image showing Mitochondria staining in Jurkat cell line. Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-Lipoamide Dehydrogenase antibody [EPR6635] (AB133551)

Immunocytochemistry/ Immunofluorescence analysis of C6 (rat glial tumor glial cell) labeling Lipoamide Dehydrogenase with ab133551 at 1/200 dilution followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed secondary antibody at 1/1000 dilution. Cells were fixed with 4% Paraformaldehyde and permeabilised with 0.1% TritonX-100. ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor® 594) at 1/200 was used as the counterstain antibody. Nuclear counter stain was DAPI. Confocal image showing Mitochondria staining in RAW 264.7 cell line. Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-Lipoamide Dehydrogenase antibody [EPR6635] (AB133551)

Immunocytochemistry/ Immunofluorescence analysis of RAW 264.7 (mouse Abelson murine leukemia virus-induced tumor macrophage) labeling Lipoamide Dehydrogenase with ab133551 at 1/200 dilution followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed secondary antibody at 1/1000 dilution. Cells were fixed with 4% Paraformaldehyde and permeabilised with 0.1% TritonX-100. ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor® 594) at 1/200 was used as the counterstain antibody. Nuclear counter stain was DAPI. Confocal image showing Mitochondria staining in RAW 264.7 cell line. Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

• WB

Lab

Western blot - Anti-Lipoamide Dehydrogenase antibody [EPR6635] (AB133551)

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

All lanes:

Western blot - Anti-Lipoamide Dehydrogenase antibody [EPR6635] (ab133551) at 1/1000 dilution

Lane 1:

Mouse brain tissue lysate at 20 µg

Lane 2:

Mouse heart tissue lysate at 20 µg

Lane 3:

Mouse kidney tissue lysate at 20 µg

Lane 4:

Mouse spleen tissue lysate at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/20000 dilution

Observed band size: 54 kDa

false

Exposure time: 1s

• WB

Unknown

Western blot - Anti-Lipoamide Dehydrogenase antibody [EPR6635] (AB133551)

Lanes 1-3 : Merged signal (red and green). Green - ab133551 observed at 56 kDa. Red - loading control ab8245 observed at 36 kDa.

ab133551 Anti-Lipoamide Dehydrogenase antibody [EPR6635] was shown to specifically react with Lipoamide Dehydrogenase in wild-type HeLa cells. Loss of signal was observed when knockout cell line ab265535 (knockout cell lysate ab257922) was used. Wild-type and Lipoamide Dehydrogenase knockout samples were subjected to SDS-PAGE. ab133551 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated overnight at 4°C at 1 in 1000 dilution and 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preadsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.

All lanes:

Western blot - Anti-Lipoamide Dehydrogenase antibody [EPR6635] (ab133551) at 1/1000 dilution

Lane 1:

Wild-type HeLa cell lysate at 20 µg

Lane 2:

DLD knockout HeLa cell lysate at 20 µg

Lane 2:

Western blot - Human DLD (Lipoamide Dehydrogenase) knockout HeLa cell line (<a href='/en-us/products/cell-lines/human-dld-lipoamide-dehydrogenase-knockout-hela-cell-line-ab265535'>ab265535</a>)

Lane 3:

Jurkat cell lysate at 20 µg

Secondary

All lanes:

Western blot - Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-irdye-800cw-preadsorbed-ab216773'>ab216773</a>) at 1/10000 dilution

Predicted band size: 54 kDa

Observed band size: 56 kDa

false

• WB

Unknown

Western blot - Anti-Lipoamide Dehydrogenase antibody [EPR6635] (AB133551)

All lanes:

Western blot - Anti-Lipoamide Dehydrogenase antibody [EPR6635] (ab133551) at 1/10000 dilution

Lane 1:

Jurkat cell lysate at 10 µg

Lane 2:

HeLa cell lysate at 10 µg

Lane 3:

293T cell lysate at 10 µg

Lane 4:

MCF7 cell lysate at 10 µg

Lane 5:

Caco-2 cell lysate at 10 µg

Secondary

All lanes:

HRP labelled goat anti-rabbit at 1/2000 dilution

Predicted band size: 54 kDa

Observed band size: 56 kDa

false

• WB

Lab

Western blot - Anti-Lipoamide Dehydrogenase antibody [EPR6635] (AB133551)

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

All lanes:

Western blot - Anti-Lipoamide Dehydrogenase antibody [EPR6635] (ab133551) at 1/1000 dilution

Lane 1:

C6 (rat glial tumor glial cell) whole cell lysate at 20 µg

Lane 2:

RAW 264.7 (mouse Abelson murine leukemia virus-induced tumor macrophage) at 20 µg

Lane 3:

PC-12 (rat adrenal gland pheochromocytoma cell) at 20 µg

Lane 4:

NIH/3T3 (mouse embryonic fibroblast) at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/20000 dilution

Observed band size: 54 kDa

false

Exposure time: 1s

• OI-RD Scanning

Unknown

OI-RD Scanning - Anti-Lipoamide Dehydrogenase antibody [EPR6635] (AB133551)

We have systematically measured KD (the equilibrium dissociation constant between the antibody and its antigen), of more than 840 recombinant antibodies to assess not only their individual KD values but also to see the average affinity of antibody. Based on the comparison with published literature values for mouse monoclonal antibodies, Recombinant antibodies appear to be on average 1-2 order of magnitude higher affinity.