Anti-DNAJA1 antibody [EPR7248]

• Flow Cyt (Intra)

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Flow Cytometry (Intracellular) - Anti-DNAJA1 antibody [EPR7248] (AB126774)

ab126774 at 1/10 dilution staining DNAJA1 in permeabilized Jurkat cells by intracellular flow cytometry (red). Rabbit IgG negative control (green).

• WB

Lab

Western blot - Anti-DNAJA1 antibody [EPR7248] (AB126774)

Lane 1 : Wild-type DNAJA1 whole cell lysate (20 μg)
Lane 2 : DNAJA1 knockout DNAJA1 whole cell lysate (20 μg)
Lane 3 : HepG2 whole cell lysate (20 μg)
Lane 4 : Jurkat whole cell lysate (20 μg)

Lanes 1 - 4 : Merged signal (red and green). Green - ab126774 observed at 45 kDa. Red - loading control, ab18058, observed at 130 kDa.

ab126774 was shown to recognize DNAJA1 in wild-type HAP1 samples along with additional cross-reactive bands. No band was observed when DNAJA1 knockout samples were examined. Wild-type and DNAJA1 knockout samples were subjected to SDS-PAGE. ab126774 and ab18058 (Mouse anti Vinculin loading control) were incubated overnight at 4°C at 1/10,000 dilution and 1/10,000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preabsorbed ab216773 and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preabsorbed ab216776 secondary antibodies at 1/10,000 dilution for 1 hour at room temperature before imaging.

All lanes:

Western blot - Anti-DNAJA1 antibody [EPR7248] (ab126774)

Predicted band size: 45 kDa

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• WB

Lab

Western blot - Anti-DNAJA1 antibody [EPR7248] (AB126774)

Lanes 1-4 : Merged signal (red and green). Green - ab126774 observed at 49 kDa. Red - loading control ab8245 observed at 36 kDa.

ab126774 Anti-DNAJA1 antibody [EPR7248] was shown to specifically react with DNAJA1 in wild-type HEK293T cells. Loss of signal was observed when knockout cell line ab266437 (knockout cell lysate ab257925) was used. Wild-type and DNAJA1 knockout samples were subjected to SDS-PAGE. ab126774 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-DNAJA1 antibody [EPR7248] (ab126774) at 1/1000 dilution

Lane 1:

Wild-type HEK293T cell lysate at 20 µg

Lane 2:

DNAJA1 knockout HEK293T cell lysate at 20 µg

Lane 2:

Western blot - Human DNAJA1 knockout HEK-293T cell line (<a href='/en-us/products/cell-lines/human-dnaja1-knockout-hek-293t-cell-line-ab266437'>ab266437</a>)

Lane 3:

HepG2 cell lysate at 20 µg

Lane 4:

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: 45 kDa

Observed band size: 49 kDa

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• WB

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Western blot - Anti-DNAJA1 antibody [EPR7248] (AB126774)

All lanes:

Western blot - Anti-DNAJA1 antibody [EPR7248] (ab126774) at 1/10000 dilution

Lane 1:

HepG2 cell lysate at 10 µg

Lane 2:

SKBR 3 cell lysate at 10 µg

Lane 3:

Jurkat cell lysate at 10 µg

Lane 4:

Raji cell lysate at 10 µg

Secondary

All lanes:

Goat anti-Rabbit HRP at 1/2000 dilution

Predicted band size: 45 kDa

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• OI-RD Scanning

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OI-RD Scanning - Anti-DNAJA1 antibody [EPR7248] (AB126774)

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.