Anti-PERP antibody [EPR7885]

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-PERP antibody [EPR7885] (AB129083)

ab129083, at a dilution of 1/100, staining PERP in A431 cells by Immunofluorescence.

• WB

Lab

Western blot - Anti-PERP antibody [EPR7885] (AB129083)

Lanes 1-3 : Merged signal (red and green). Green - ab129083 observed at 24 kDa. Red - loading control ab8245 observed at 36 kDa.

ab129083 Anti-PERP antibody [EPR7885] was shown to specifically react with PERP in wild-type HeLa cells. Loss of signal was observed when knockout cell line ab265335 (knockout cell lysate ab258105) was used. Wild-type and PERP knockout samples were subjected to SDS-PAGE. ab129083 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated at room temperature for 2.5 hours 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-PERP antibody [EPR7885] (ab129083) at 1/1000 dilution

Lane 1:

Wild-type HeLa cell lysate at 20 µg

Lane 2:

PERP knockout HeLa cell lysate at 20 µg

Lane 2:

Western blot - Human PERP knockout HeLa cell line (<a href='/en-us/products/cell-lines/human-perp-knockout-hela-cell-line-ab265335'>ab265335</a>)

Lane 3:

HaCaT 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: 21 kDa

Observed band size: 24 kDa

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• WB

Lab

Western blot - Anti-PERP antibody [EPR7885] (AB129083)

False colour image of Western blot : Anti-PERP antibody [EPR7885] staining at 1/1000 dilution, shown in green; Mouse anti-Alpha Tubulin [DM1A] (ab7291) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab129083 was shown to bind specifically to PERP. A band was observed at 17 kDa in wild-type A431 cell lysates with no signal observed at this size in PERP knockout cell line ab270475 (knockout cell lysate ab270498). To generate this image, wild-type and PERP knockout A431 cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3% milk in TBS-0.1% Tween® 20 (TBS-T) before incubation with primary antibodies overnight at 4°C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (ab216776) at 1/20000 dilution.

All lanes:

Western blot - Anti-PERP antibody [EPR7885] (ab129083) at 1/1000 dilution

Lane 1:

Wild-type A431 cell lysate at 20 µg

Lane 2:

PERP knockout A431 cell lysate at 20 µg

Lane 3:

HeLa cell lysate at 20 µg

Lane 4:

MOLT-4 cell lysate at 20 µg

Predicted band size: 21 kDa

Observed band size: 17 kDa

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• WB

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Western blot - Anti-PERP antibody [EPR7885] (AB129083)

All lanes:

Western blot - Anti-PERP antibody [EPR7885] (ab129083) at 1/1000 dilution

Lane 1:

Mouse heart lysate at 10 µg

Lane 2:

Rat heart lysate at 10 µg

Lane 3:

A431 lysate at 10 µg

Lane 4:

HACAT lysate at 10 µg

Lane 5:

Human skin lysate at 10 µg

Secondary

All lanes:

HRP labelled goat anti-rabbit at 1/2000 dilution

Predicted band size: 21 kDa

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• OI-RD Scanning

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OI-RD Scanning - Anti-PERP antibody [EPR7885] (AB129083)

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.