Recombinant Anti-Moesin antibody [EPR2428(2)] (ab151542)

False colour image of Western blot: Anti-Moesin antibody [EPR2428(2)] staining at 2 µg/ml, shown in green; Mouse anti-GAPDH antibody [6C5] (ab8245) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab151542 was shown to bind specifically to Moesin. A band was observed at 75 kDa in wild-type HeLa cell lysates with no signal observed at this size in MSN knockout cell line ab265020 (knockout cell lysate ab257542). To generate this image, wild-type and MSN knockout HeLa 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.

Lane 1: Wild-type HAP1 whole cell lysate (20 µg)
Lane 2: Moesin knockout HAP1 whole cell lysate (20 µg)
Lane 3: HeLa whole cell lysate (20 µg)
Lane 4: Raji whole cell lysate (20 µg)

Lanes 1 - 4: Merged signal (red and green). Green - ab151542 observed at 75 kDa. Red - loading control, ab9484, observed at 37 kDa.

ab151542 was shown to specifically react with Moesin in wild-type HAP1 cells as signal was lost in Moesin knockout cells. Wild-type and Moesin knockout samples were subjected to SDS-PAGE. Ab151542 and ab9484 (Mouse anti-GAPDH loading control) were incubated overnight at 4°C at 1/1000 dilution and 1/20000 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 1/20000 dilution for 1 hour at room temperature before imaging.

ab151542 was shown to react with MSN in wild-type HeLa cells in Immunocytochemistry with loss of signal observed in MSN knockout cell line ab265020. Wild-type and knockout cells were mixed and pelleted at a 1:1 ratio on coverslips. The cells were fixed with 4% paraformaldehyde (15 min) then permeabilized with 0.1% Triton X-100 (10min) and then blocked with 1/10000. The cells were then incubated with ab151542 at 1/200 dilution overnight at 4°C followed by a further incubation at room temperature for 1h with a goat anti-rabbit secondary antibody to (Alexa Fluor^® 555) at 0.5 μg/ml. Acquisition of the green (wild-type), red (antibody staining) and far-red (knockout) channels was performed. Representative grayscale images of the red channel are shown. Wild-type and knockout cells are outlined with yellow and magenta dashed line, respectively. Schematic representation of the mosaic strategy used is shown on the bottom-right panel. Image was acquired with a Zeiss(LSM-880). These data were provided by YCharOS Inc., an open science company with the mission of characterizing commercially available antibody reagents for all human proteins. Abcam and YCharOS are working together to help address the reproducibility crisis by enabling the life science community to better evaluate commercially available antibodies.

ab151542 was shown to react with MSN in wild-type HeLa cells in Western blot with loss of signal observed in MSN knockout cell line ab265020 (MSN knockout cell lysate ab257542). Wild-type HeLa and MSN knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in 5% milk in TBST for 1 hr before incubation with ab151542 overnight at 4 °C at a 1/1000 dilution. Blots were incubated with goat anti-rabbit HRP secondary antibodies at 0.2ug/mL before imaging. These data were provided by YCharOS Inc., an open science company with the mission of characterizing commercially available antibody reagents for all human proteins. Abcam and YCharOS are working together to help address the reproducibility crisis by enabling the life science community to better evaluate commercially available antibodies.

Immunohistochemical analysis of paraffin-embedded Human breast carcinoma tissue labeling Moesin with ab151542 at 1/100 dilution.

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