Anti-Vinculin antibody [EPR8185] - Loading Control

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-Vinculin antibody [EPR8185] - Loading Control (AB129002)

Immunofluorescent staining of HEK293 cells (fixed in 4% PFA, permeabilized with 0.1% Triton X 100) using purified ab129002 at a dilution of 1/50. An Alexa Fluor^® 488 goat anti-rabbit antibody (ab150077) was used as the secondary at a dilution of 1/1000 and the cells were counter stained with DAPI. The negative controls are shown in the bottom middle and right hand panels. For negative control 1, the primary was used and then goat anti-mouse IgG was used at a dilution of 1/500. For negative control 2, a mouse primary antibody (ab7291) and anti-rabbit secondary antibody (ab150077) were used.

• WB

Lab

Western blot - Anti-Vinculin antibody [EPR8185] - Loading Control (AB129002)

Western blot : Anti-VCL antibody [EPR8185] (ab129002) staining at 1/10000 dilution, shown in green; Mouse anti-CANX [CANX/1543] (ab238078) loading control staining at 1/20000 dilution, shown in magenta. In Western blot, ab129002 was shown to bind specifically to VCL. A band was observed at 124 kDa in wild-type A431 cell lysates with no signal observed at this size in VCL knockout cell line. To generate this image, wild-type and VCL 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 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution.

All lanes:

Western blot - Anti-Vinculin antibody [EPR8185] - Loading Control (ab129002) at 1/10000 dilution

Lane 1:

Wild-type A431 cell lysate

Lane 2:

Western blot - Human VCL (Vinculin) knockout A-431 cell line (<a href='/en-us/products/cell-lines/human-vcl-vinculin-knockout-a-431-cell-line-ab261898'>ab261898</a>) at 20 µg

Lane 2:

VCL knockout A431 cell lysate

Lane 3:

HeLa cell lysate

Lane 4:

Jurkat cell lysate

false

• ICC/IF

Unknown

Immunocytochemistry/ Immunofluorescence - Anti-Vinculin antibody [EPR8185] - Loading Control (AB129002)

Immunofluorescent staining of vinculin in HUVEC cells with unpurified ab129002 at 1/100 dilution.

• Flow Cyt (Intra)

Unknown

Flow Cytometry (Intracellular) - Anti-Vinculin antibody [EPR8185] - Loading Control (AB129002)

Intracellular Flow Cytometry analysis of 293 (human embryonic kidney epithelial) cells labeling Vinculin (red) with ab129002 at a 1/200 dilution. Cells were fixed with 4% paraformaldehyde and permeabilized with 90% methanol. A goat anti-rabbit IgG (Alexa Fluorr^® 488) (ab150077) was used as the secondary antibody at a 1/2000 dilution. Black - Rabbit monoclonal IgG (ab172730). Blue (unlabeled control) - Cells without incubation with the primary and secondary antibodies.

• Flow Cyt (Intra)

Lab

Flow Cytometry (Intracellular) - Anti-Vinculin antibody [EPR8185] - Loading Control (AB129002)

Flow cytometry overlay histogram showing wild-type A-431 (green line) and VCL knockout A-431 stained with ab129002 (red line). The cells were fixed with 80% methanol (5 min) and then permeabilised with 0.1% PBS-Triton X-100 for 15 min. The cells were then incubated in 1x PBS containing 10% normal goat serum to block non-specific protein-protein interaction followed by the antibody (ab129002) (1x 106 in 100μl at 0.00032 μg/ml (1/6249999)) for 30min at 22°C.

The secondary antibody Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed was incubated at 1/4000 for 30min at 22°C

Isotype control antibody Recombinant Rabbit IgG, monoclonal [EPR25A] - Isotype Control was used at the same concentration and conditions as the primary antibody (wild-type A-431 - black line, VCL knockout A-431 - grey line). Unlabelled sample was also used as a control (this line is not shown for the purpose of simplicity).

Acquisition of >5000 events were collected using a 50 mW Blue laser (488nm) and 525/40 bandpass filter.

• Flow Cyt (Intra)

Lab

Flow Cytometry (Intracellular) - Anti-Vinculin antibody [EPR8185] - Loading Control (AB129002)

Flow cytometry overlay histogram showing left HeLa positive cells and right negative HeLa VCL KO stained with ab129002 (red line). The cells were fixed with 80% methanol (5 min) and then permeabilised with 0.1% PBS-Triton X-100 for 15 min. The cells were then incubated in 1x PBS containing 10% normal goat serum to block non-specific protein-protein interaction followed by the antibody (ab129002) (1x 10⁶ in 100μl at 0.008μg/ml (1/7875)) for 30min at 22°C.

The secondary antibody Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed was incubated at 1/4000 for 30min at 22°C

Isotype control antibody (black line) was Recombinant Rabbit IgG, monoclonal [EPR25A] - Isotype Control used at the same concentration and conditions as the primary antibody. Unlabelled sample (blue line) was also used as a control.

Acquisition of >5000 events were collected using a 50 mW Blue laser (488nm) and 525/40 bandpass filter.

• IP

Lab

Immunoprecipitation - Anti-Vinculin antibody [EPR8185] - Loading Control (AB129002)

ab129002 (purified) at 1/20 immunoprecipitating vinculin in HeLa cells. Lane 1 : HeLa whole cell lysate (10 μg). Lane 2 : HeLa whole cell lysate (10 μg). Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab129002 in HeLa whole cell lysate. For western blotting, a HRP-conjugated goat anti-rabbit antibody was used as the secondary antibody (1/1000).

Blocking buffer and concentration : 5% NFDM/TBST.

Diluting buffer and concentration : 5% NFDM /TBST.

All lanes:

Immunoprecipitation - Anti-Vinculin antibody [EPR8185] - Loading Control (ab129002)

Predicted band size: 124 kDa

false

• WB

Supplier Data

Western blot - Anti-Vinculin antibody [EPR8185] - Loading Control (AB129002)

In Western blot, Anti-Vinculin antibody [EPR8185] (ab129002) staining at 1/10000 dilution.

All lanes:

Western blot - Anti-Wnt11 antibody [EPR25633-141] (<a href='/en-us/products/primary-antibodies/wnt11-antibody-epr25633-141-ab316738'>ab316738</a>) at 1/1000 dilution

Lane 1:

Untreated JEG-3 (human placenta epithelial cell) whole cell lysate at 60 µg

Lane 2:

JEG-3 treated with 300 ng/ml Brefeldin A for 24h whole cell lysate at 60 µg

Secondary

All lanes:

Goat Anti-Rabbit IgG (HRP) with minimal cross-reactivity with human IgG at 1/2000 dilution

Observed band size: 39 kDa,124 kDa

false

Exposure time: 8s

• WB

Supplier Data

Western blot - Anti-Vinculin antibody [EPR8185] - Loading Control (AB129002)

Negative control : T-47D, MCF7.

Samples are non-boiled as boiling may cause protein aggregation.

In Western blot, Anti-Vinculin antibody [EPR8185] (ab129002) staining at 1/10000 dilution.

All lanes:

Western blot - Anti-PIEZO2 antibody [EPR27152-48] (<a href='/en-us/products/primary-antibodies/piezo2-antibody-epr27152-48-ab318187'>ab318187</a>) at 1/1000 dilution

Lane 1:

MKL-1 (human Skin Loosely packed floati aggregates with irregular outline and no central necrosis) whole cell lysate at 48 µg

Lane 2:

T-47D (human ductal breast epithelial tumor epithelial cell) whole cell lysate at 48 µg

Lane 3:

MCF7 (human breast adenocarcinoma epithelial cell) whole cell lysate at 48 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: 320 kDa,124 kDa

false

Exposure time: 92s

• WB

Supplier Data

Western blot - Anti-Vinculin antibody [EPR8185] - Loading Control (AB129002)

Negative control : NCI-H1975 (PMID : 31231131), A549 (PMID : 31231131).

In Western blot, Anti-GAPDH antibody [EPR16891] - Loading Control (ab181602) staining at 1/200000 dilution.

In Western blot, Anti-Vinculin antibody [EPR8185] (ab129002) staining at 1/10000 dilution.

Exposure time : Lane 1-2 : 3 seconds, Lane 3-4 : 26 seconds

All lanes:

Western blot - Anti-Wnt11 antibody [EPR25633-141] (<a href='/en-us/products/primary-antibodies/wnt11-antibody-epr25633-141-ab316738'>ab316738</a>) at 1/1000 dilution

Lane 1:

TF-1 (human Erythroleukemia erythroblast) whole cell lysate at 60 µg

Lane 2:

NCI-H1975 (human adenocarcinoma lung epithelial cell) whole cell lysate at 60 µg

Lane 3:

Saos-2 (human osteosarcoma epithelial cell) whole cell lysate at 60 µg

Lane 4:

A549 (human lung carcinoma epithelial cell) whole cell lysate at 60 µg

Secondary

All lanes:

Goat Anti-Rabbit IgG (HRP) with minimal cross-reactivity with human IgG at 1/2000 dilution

Observed band size: 39 kDa,36 kDa,124 kDa

false

• WB

Supplier Data

Western blot - Anti-Vinculin antibody [EPR8185] - Loading Control (AB129002)

Lysates were freshly made and used for Western blotting immediately to minimize protein degradation.

In Western blot, Anti-Vinculin antibody [EPR8185] (ab129002) staining at 1/10000 dilution.

All lanes:

Western blot - Anti-CSB antibody [EPR27019-44] (<a href='/en-us/products/primary-antibodies/csb-antibody-epr27019-44-ab316744'>ab316744</a>) at 1/1000 dilution

Lane 1:

U-2OS (human bone osteosarcoma epithelial cell) whole cell lysate at 20 µg

Lane 2:

MCF7 (human breast adenocarcinoma epithelial cell) whole cell 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 1/100000 dilution

Observed band size: 200 kDa,124 kDa

false

Exposure time: 180s

• WB

Lab

Western blot - Anti-Vinculin antibody [EPR8185] - Loading Control (AB129002)

Blocking and diluting buffer and concentration : 5% NFDM/TBST.

The expression profile/ molecular weight observed is consistent with what has been described in the literature (PMID : 22558357; PMID : 19166587).

To minimize protein degradation, cells were lysed immediately after harvest and then applied to a gel and transfer membrane for Western blotting as soon as possible.

The identity of the lower band at approximately 140 kDa is likely SET-NUP214 fusion protein.

In Western blot, Anti-Vinculin antibody [EPR8185] (ab129002) (1 : 10000) (124KDa).

Exposure time : Lanes 1-2 : 180 seconds; Lanes 3 : 37 seconds

All lanes:

Western blot - Anti-NUP214 antibody [EPR29100-623] (<a href='/en-us/products/primary-antibodies/nup214-antibody-epr29100-623-ab325238'>ab325238</a>) at 1/1000 dilution

Lane 1:

HeLa (human cervical adenocarcinoma epithelial cell) whole cell lysate at 20 µg

Lane 2:

293T (human embryonic kidney epithelial cell) whole cell lysate at 20 µg

Lane 3:

K-562 (human chronic myelogenous leukemia lymphoblast) whole cell 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: 268 kDa,124 kDa

false

• WB

Lab

Western blot - Anti-Vinculin antibody [EPR8185] - Loading Control (AB129002)

Blocking and diluting buffer and concentration : 5% NFDM/TBST.

Performed under reducing conditions.

In Western blot, ab325390 was shown to bind specifically to ATRX. Target of interest was observed at 310 kDa in wild-type A549 cell lysates (lane 1) with no signal observed at this size in ATRX knockout cell line (lane 2, knockout cell line ab287232).

To minimize protein degradation, cells were lysed immediately after harvest and then applied to a gel and transfer membrane for Western blotting as soon as possible.

In Western blot, Anti-Vinculin antibody [EPR8185] (ab129002) (1 : 10000) (124KDa).

All lanes:

Western blot - Anti-ATRX antibody [EPR29209-280] (<a href='/en-us/products/primary-antibodies/atrx-antibody-epr29209-280-ab325390'>ab325390</a>) at 1/1000 dilution

Lane 1:

Wild-type A549 (human lung carcinoma epithelial cell) whole cell lysate at 20 µg

Lane 2:

ATRX knockout A549 whole cell lysate

Lane 3:

Wild-type A549 whole cell 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 1/100000 dilution

Observed band size: 310 kDa,124 kDa

false

Exposure time: 180s

• WB

Lab

Western blot - Anti-Vinculin antibody [EPR8185] - Loading Control (AB129002)

Blocking and diluting buffer and concentration : 5% NFDM/TBST.

Negative control : adult skeletal muscle, adult lung (PMID : 25957469; PMID : 3530499).

Low expression : HEK-293 (PMID : 25957469)

The expression profile observed is consistent with what has been described in the literature (PMID : 3530499, PMID : 12899730, PMID : 31579487, PMID : 25957469 ).

In Western blot, Anti-Vinculin antibody [EPR8185] (ab129002) (1 : 10000) (124KDa).

Exposure time : Lanes 1-4 : 59 seconds; Lanes 5-9 : 15 seconds

All lanes:

Western blot - Anti-heavy chain Myosin/MYH3 antibody [EPR29772-555] (<a href='/en-us/products/primary-antibodies/heavy-chain-myosin-myh3-antibody-epr29772-555-ab325795'>ab325795</a>) at 1/1000 dilution

Lane 1:

Untreated C2C12 (mouse myoblast) whole cell lysate at 20 µg

Lane 2:

C2C12 differentiation to muscle for 3 days whole cell lysate at 20 µg

Lane 3:

RD (human muscle multinucleated spindle-shaped cell) whole cell lysate at 20 µg

Lane 4:

HEK-293 (human embryonic kidney epithelial cell) whole cell lysate at 20 µg

Lane 5:

Mouse E14 forelimb tissue lysate at 20 µg

Lane 6:

Mouse embryo tissue lysate at 20 µg

Lane 7:

Mouse E12.5 embryo tissue lysate at 20 µg

Lane 8:

Mouse adult skeletal muscle tissue lysate at 20 µg

Lane 9:

Mouse adult lung 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: 223 kDa,124 kDa

false

• WB

Supplier Data

Western blot - Anti-Vinculin antibody [EPR8185] - Loading Control (AB129002)

The identity of the higher MW band at approximately 250 kDa is unknown.

In Western blot, Anti-Vinculin antibody [EPR8185] (ab129002) staining at 1/10000 dilution.

All lanes:

Western blot - Anti-LARP1 antibody [EPR28797-92] (<a href='/en-us/products/primary-antibodies/larp1-antibody-epr28797-92-ab317575'>ab317575</a>) at 1/1000 dilution

Lane 1:

HeLa (human cervical adenocarcinoma epithelial cell) transfected with scrambled siRNA control whole cell lysate at 20 µg

Lane 2:

HeLa transfected with siRNA specifically targeting LARP1 whole cell lysate at 20 µg

Lane 3:

PC-3 (human prostate adenocarcinoma epithelial cell) whole cell lysate at 20 µg

Lane 4:

SW480 (human colorectal adenocarcinoma epithelial cell) whole cell lysate at 20 µg

Lane 5:

HEK293T (human embryonic kidney epithelial cell) whole cell 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: 150 kDa,124 kDa

false

Exposure time: 15s

• WB

Supplier Data

Western blot - Anti-Vinculin antibody [EPR8185] - Loading Control (AB129002)

Negative control : HEL

In Western blot, Anti-Vinculin antibody [EPR8185] (ab129002) staining at 1/10000 dilution.

All lanes:

Western blot - Anti-Agrin antibody [EPR28043-55] (<a href='/en-us/products/primary-antibodies/agrin-antibody-epr28043-55-ab316754'>ab316754</a>) at 1/1000 dilution

Lane 1:

HaCaT (human skin keratinocyte) whole cell lysate at 40 µg

Lane 2:

HEL (human Erythroleukemia erythroblast) whole cell lysate at 40 µg

Lane 3:

Huh7 (human hepatocellar carcinoma epithelial cell) whole cell lysate at 40 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: 260 kDa,124 kDa

false

Exposure time: 180s

• WB

Supplier Data

Western blot - Anti-Vinculin antibody [EPR8185] - Loading Control (AB129002)

This antibody was tested on fresh and frozen HeLa and 293T Whole Cell Lysate. The Lysates in lanes 3-4 were freshly made and used for Western Blotting immediately to minimize protein degradation.

In Western blot, Anti-Vinculin antibody [EPR8185] (ab129002) staining at 1/10000 dilution.

Exposure time : Lane 1-2 : 103 seconds; Lane 3-4 : 180 seconds

All lanes:

Western blot - Anti-CSB antibody [EPR27019-44] (<a href='/en-us/products/primary-antibodies/csb-antibody-epr27019-44-ab316744'>ab316744</a>) at 1/1000 dilution

Lane 1:

HeLa (human cervical adenocarcinoma epithelial cell) whole cell lysate at 20 µg

Lane 2:

293T (human embryonic kidney epithelial cell) whole cell lysate at 20 µg

Lane 3:

HeLa whole cell lysate at 20 µg

Lane 4:

293T whole cell 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 1/100000 dilution

Observed band size: 200 kDa,124 kDa

false

• WB

Supplier Data

Western blot - Anti-Vinculin antibody [EPR8185] - Loading Control (AB129002)

The identity of the band at 100 kDa is unknown.

The bands beneath the target band between 100 kDa and 260 kDa (Lane 1-8) are likely to be degraded target fragments.

In Western blot, Anti-Vinculin antibody [EPR8185] (ab129002) staining at 1/10000 dilution.

Exposure time : Lane 1-8 : 15 seconds; Lane 9-12 : 48 seconds

All lanes:

Western blot - Anti-POLE antibody [EPR28639-64] (<a href='/en-us/products/primary-antibodies/pole-antibody-epr28639-64-ab318968'>ab318968</a>) at 1/1000 dilution

Lane 1:

HeLa (human cervical adenocarcinoma epithelial cell) whole cell lysate at 20 µg

Lane 2:

293T (human embryonic kidney epithelial cell) whole cell lysate at 20 µg

Lane 3:

MCF7 (human breast adenocarcinoma epithelial cell) whole cell lysate at 20 µg

Lane 4:

THP-1 (human monocytic leukemia monocyte) whole cell lysate at 20 µg

Lane 5:

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

Lane 6:

F9 (mouse embryonal carcinoma epithelial cell) whole cell lysate at 20 µg

Lane 7:

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

Lane 8:

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

Lane 9:

HeLa (human cervical adenocarcinoma epithelial cell) whole cell fresh lysate at 20 µg

Lane 10:

293T (human embryonic kidney epithelial cell) whole cell fresh lysate at 20 µg

Lane 11:

NIH/3T3 (mouse embryonic fibroblast) whole cell fresh lysate at 20 µg

Lane 12:

PC-12 (rat adrenal gland pheochromocytoma cell) whole cell fresh 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 1/100000 dilution

Observed band size: 260 kDa,124 kDa

false

• WB

Lab

Western blot - Anti-Vinculin antibody [EPR8185] - Loading Control (AB129002)

Blocking and diluting buffer and concentration : 5% NFDM/TBST.

The expression profile/ molecular weight observed is consistent with what has been described in the literature (PMID : 22558357; PMID : 19166587).

To minimize protein degradation, cells were lysed immediately after harvest and then applied to a gel and transfer membrane for Western blotting as soon as possible.

The identity of the lower band at approximately 140 kDa is likely a SET-NUP214 fusion protein.

In Western blot, Anti-Vinculin antibody [EPR8185] (ab129002) 1 : 10000 (124Kda).

All lanes:

Western blot - Anti-NUP214 antibody [EPR29100-623] (<a href='/en-us/products/primary-antibodies/nup214-antibody-epr29100-623-ab325238'>ab325238</a>) at 1/1000 dilution

Lane 1:

HeLa (human cervical adenocarcinoma epithelial cell) transfected with scrambled siRNA control whole cell lysate at 48 µg

Lane 2:

HeLa transfected with siRNA specifically targeting NUP214 whole cell lysate at 48 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: 268 kDa,124 kDa

false

Exposure time: 180s

• WB

Lab

Western blot - Anti-Vinculin antibody [EPR8185] - Loading Control (AB129002)

Blocking and diluting buffer and concentration : 5% NFDM/TBST.

This blot was developed using a high-sensitivity ECL substrate, allowing for the detection of proteins in the mid-femtogram range.

To minimize protein degradation, the fresh lysates were lysed immediately after harvest and then applied to a gel and transfer membrane for Western blotting as soon as possible.

In Western blot, Anti-Vinculin antibody [EPR8185] (ab129002) (1 : 10000) (124KDa).

All lanes:

Western blot - Anti-ATR antibody [EPR30640-130] (<a href='/en-us/products/primary-antibodies/atr-antibody-epr30640-130-ab325605'>ab325605</a>) at 1/1000 dilution

Lane 1:

HeLa (human cervical adenocarcinoma epithelial cell) transfected with scrambled siRNA control whole cell lysate at 20 µg

Lane 2:

HeLa transfected with siRNA specifically targeting ATR whole cell 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: 270 kDa,124 kDa

true

Exposure time: 180s

• WB

Supplier Data

Western blot - Anti-Vinculin antibody [EPR8185] - Loading Control (AB129002)

The identity of the band at 100 kDa is unknown.

The bands beneath the target band between 100 kDa and 260 kDa are likely to be degraded target fragments.

In Western blot, ab318968 was shown to bind specifically to POLE. Target of interest was observed at 260kDa in wild-type HAP1 cell lysates (Lane 1) with no signal observed at this size in POLE knockout cell line (Lane 2).

In Western blot, Anti-Vinculin antibody [EPR8185] (ab129002) staining at 1/10000 dilution.

All lanes:

Western blot - Anti-POLE antibody [EPR28639-64] (<a href='/en-us/products/primary-antibodies/pole-antibody-epr28639-64-ab318968'>ab318968</a>) at 1/1000 dilution

Lane 1:

parental HAP1(HOWT01) (human chronic myelogenous leukemia near-haploid cell ) whole cell lysate at 20 µg

Lane 2:

POLE Knockout HAP1(HO010404) whole cell 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 1/100000 dilution

Observed band size: 260 kDa,124 kDa

false

Exposure time: 180s

• WB

Supplier Data

Western blot - Anti-Vinculin antibody [EPR8185] - Loading Control (AB129002)

Negative control : kidney

In Western blot, Anti-Vinculin antibody [EPR8185] (ab129002) staining at 1/10000 dilution.

Exposure time : Lanes 1-2 : 10 seconds, Lanes 3-8 : 6 seconds

Blocking and diluting buffer and concentration : 5% NFDM/TBST.

All lanes:

Western blot - Anti-NEFH antibody [EPR20020] - Neuronal Marker - Chicken IgY (Chimeric) (<a href='/en-us/products/primary-antibodies/nefh-antibody-epr20020-neuronal-marker-chicken-igy-chimeric-ab317042'>ab317042</a>) at 1/1000 dilution

Lane 1:

Human Brainstem tissue lysate at 20 µg

Lane 2:

Human kidney tissue lysate at 20 µg

Lane 3:

Mouse cerebellum tissue lysate at 20 µg

Lane 4:

Mouse brain tissue lysate at 20 µg

Lane 5:

Mouse kidney tissue lysate at 20 µg

Lane 6:

Rat cerebellum tissue lysate at 20 µg

Lane 7:

Rat brain tissue lysate at 20 µg

Lane 8:

Rat kidney tissue lysate at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Chicken IgY H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-chicken-igy-h-l-hrp-ab6877'>ab6877</a>) at 1/100000 dilution

Observed band size: 200-210 kDa,124 kDa

false

• WB

Supplier Data

Western blot - Anti-Vinculin antibody [EPR8185] - Loading Control (AB129002)

In Western blot, Anti-Vinculin antibody [EPR8185] (ab129002) staining at 1/10000 dilution.

All lanes:

Western blot - Anti-UNG2 antibody [EPR29004-78] (<a href='/en-us/products/primary-antibodies/ung2-antibody-epr29004-78-ab318982'>ab318982</a>) at 1/1000 dilution

Lane 1:

HeLa (human cervical adenocarcinoma epithelial cell) transfected with scrambled siRNA control whole cell lysate at 50 µg

Lane 2:

HeLa transfected with siRNA specifically targeting UNG2 whole cell lysate at 50 µg

Lane 3:

293T (human embryonic kidney epithelial cell) whole cell lysate at 50 µg

Lane 4:

Caco-2 (human colorectal adenocarcinoma epithelial cell) whole cell lysate at 50 µg

Lane 5:

T-47D (human ductal breast epithelial tumor epithelial cell) whole cell lysate at 50 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 1/100000 dilution

Observed band size: 37 kDa,124 kDa

false

Exposure time: 180s

• WB

Supplier Data

Western blot - Anti-Vinculin antibody [EPR8185] - Loading Control (AB129002)

The expression profile/ molecular weight observed isconsistent with what has been described in the literature (PMID : 34736895).

Lysates were freshly made and used for Western blotting immediately to minimize protein degradation.

The identity of the band below 100 kDa is unknown.

In Western blot, Anti-Vinculin antibody [EPR8185] (ab129002) staining at 1/10000 dilution.

All lanes:

Western blot - Anti-Rif1 antibody [EPR27131-83] (<a href='/en-us/products/primary-antibodies/rif1-antibody-epr27131-83-ab316748'>ab316748</a>) at 1/1000 dilution

Lane 1:

HeLa (human cervical adenocarcinoma epithelial cell) transfected with scrambled siRNA control whole cell lysate at 40 µg

Lane 2:

HeLa transfected with siRNA specifically targeting Rif1 whole cell lysate at 40 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: 110-310 kDa,124 kDa

false

Exposure time: 180s

• WB

Supplier Data

Western blot - Anti-Vinculin antibody [EPR8185] - Loading Control (AB129002)

Blocking and diluting buffer and concentration : 5% NFDM/TBST.

The expression of CCL1 is upregulated in response to PMA, GM-CSF and BFA treatment.

In Western blot, Anti-Vinculin antibody [EPR8185] (ab129002) staining at 1/10000 dilution.

All lanes:

Western blot - Anti-CCL1 antibody [EPR28766-53] (<a href='/en-us/products/primary-antibodies/ccl1-antibody-epr28766-53-ab317571'>ab317571</a>) at 1/1000 dilution

Lane 1:

U-937 (human histiocytic lymphoma monocyte) treated with 100nM PMA for 3 days whole cell lysate at 60 µg

Lane 2:

U-937 treated first with 100nM PMA for 3 days, then replaced with 50ng/ml GM-CSF for 24h, 5µg/ml BFA was then added for additional 4h at 60 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: 15 kDa,124 kDa

false

Exposure time: 114s

• WB

Supplier Data

Western blot - Anti-Vinculin antibody [EPR8185] - Loading Control (AB129002)

Negative control : HeLa, Raji (PMID : 35839778), human skeletal muscle.

In Western blot, Anti-Vinculin antibody [EPR8185] (ab129002) staining at 1/10000 dilution.

Exposure time : Lanes 1-4, 6-7 : 10 seconds; Lanes 5 : 1 second;

All lanes:

Western blot - Anti-Lrp2 / Megalin antibody [RM1150] (<a href='/en-us/products/primary-antibodies/lrp2-megalin-antibody-rm1150-ab317333'>ab317333</a>) at 1/1000 dilution

Lane 1:

JEG-3 (human placenta epithelial cell) whole cell lysate at 40 µg

Lane 2:

HeLa (human cervical adenocarcinoma epithelial cell) whole cell lysate at 40 µg

Lane 3:

Raji (human Burkitts lymphoma B lymphocyte) whole cell lysate at 40 µg

Lane 4:

Caco-2 (human colorectal adenocarcinoma epithelial cell) whole cell lysate at 40 µg

Lane 5:

Human kidney tissue lysate at 40 µg

Lane 6:

Human placenta tissue lysate at 40 µg

Lane 7:

Human skeletal muscle tissue lysate at 40 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: 522 kDa,124 kDa

false

• WB

Lab

Western blot - Anti-Vinculin antibody [EPR8185] - Loading Control (AB129002)

Blocking and diluting buffer and concentration : 5% NFDM/TBST. ab129002 was used as a loading control.

All lanes:

Western blot - Anti-Creatine Kinase MM antibody [3E5BA5] (<a href='/en-us/products/primary-antibodies/creatine-kinase-mm-antibody-3e5ba5-ab110319'>ab110319</a>) at 1/50 dilution

Lane 1:

Mouse skeletal muscle tissue lysate at 20 µg

Lane 2:

Mouse spleen tissue lysate at 20 µg

Lane 3:

Mouse placenta tissue lysate at 20 µg

Lane 4:

Human skeletal muscle tissue lysate at 20 µg

Lane 5:

Human spleen tissue lysate at 20 µg

Lane 6:

Human placenta tissue lysate at 20 µg

Predicted band size: 43 kDa

Observed band size: 43 kDa

false

Exposure time: 180s

• WB

Supplier Data

Western blot - Anti-Vinculin antibody [EPR8185] - Loading Control (AB129002)

The identity of the bands between 100 kDa and 130 kDa are unknown.

In Western blot, Anti-Vinculin antibody [EPR8185] (ab129002) staining at 1/10000 dilution.

In Western blot, Anti-ATR antibody - Total protein control staining at 1/1000 dilution.

All lanes:

Western blot - Anti-ATR (phospho S428) antibody [EPR25772-3F09] (<a href='/en-us/products/primary-antibodies/atr-phospho-s428-antibody-epr25772-3f09-ab316925'>ab316925</a>) at 1/1000 dilution

Lane 1:

Untreated HT-29 (human colorectal adenocarcinoma epithelial cell) whole cell lysate (untreated membrane) at 20 µg

Lane 2:

HT-29 treated with 100mJ/cm2 UV, then recovery for 30 minutes, whole cell lysate (untreated membrane) at 20 µg

Lane 3:

HT-29 treated with 100mJ/cm2 UV, then recovery for 30 minutes, whole cell lysate (Lambda phosphatase treated membrane) 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: 290 kDa,124 kDa

true

Exposure time: 180s

• WB

Supplier Data

Western blot - Anti-Vinculin antibody [EPR8185] - Loading Control (AB129002)

Blocking and diluting buffer and concentration : 5% NFDM/TBST.

Low expression : PC-3

In Western blot, Anti-Vinculin antibody [EPR8185] (ab129002) staining at 1/10000 dilution.

All lanes:

Western blot - Anti-RASA2 antibody [EPR29503-39] (<a href='/en-us/products/primary-antibodies/rasa2-antibody-epr29503-39-ab323554'>ab323554</a>) at 1/1000 dilution

Lane 1:

DLD-1 (human colorectal adenocarcinoma epithelial cell) whole lysate at 20 µg

Lane 2:

PC-3 (human prostate adenocarcinoma epithelial cell) whole lysate at 20 µg

Secondary

All lanes:

Goat Anti-Rabbit IgG (HRP) with minimal cross-reactivity with human IgG at 1/2000 dilution

Observed band size: 96 kDa,124 kDa

false

Exposure time: 180s

• WB

Supplier Data

Western blot - Anti-Vinculin antibody [EPR8185] - Loading Control (AB129002)

Blocking and diluting buffer and concentration : 5% NFDM/TBST.

Negative control : HepG2, K-562

Samples are non-boiled as boiling may cause protein aggregation.

In Western blot, Anti-Vinculin antibody [EPR8185] (ab129002) staining at 1/10000 dilution.

All lanes:

Western blot - Anti-GPR177/WLS antibody [EPR29686-569] (<a href='/en-us/products/primary-antibodies/gpr177-wls-antibody-epr29686-569-ab323481'>ab323481</a>) at 1/1000 dilution

Lane 1:

HaCaT (human skin keratinocyte) whole lysate at 50 µg

Lane 2:

A-204 (human muscle rhabdomyosarcoma cell) whole lysate at 50 µg

Lane 3:

HepG2 (human hepatocellular carcinoma epithelial cell) whole lysate at 50 µg

Lane 4:

K-562 (human chronic myelogenous leukemia lymphoblast) whole lysate at 50 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: 45 kDa,124 kDa

false

Exposure time: 26s

• WB

Supplier Data

Western blot - Anti-Vinculin antibody [EPR8185] - Loading Control (AB129002)

Blocking and diluting buffer and concentration : 5% NFDM/TBST.

Low expression : CT26.WT, U-937.

The expression profile/ molecular weight observed is consistent with what has been described in the literature (PMID : 20944749, 17925379, 19692571, 25239622).

Samples are non-boiled as boiling may cause protein aggregation.

In Western blot, Anti-Vinculin antibody [EPR8185] (ab129002) (124 kDa) staining at 1/10000 dilution.

Exposure time : Lanes 1-2 : 59 seconds, lanes 3-4 : 26 seconds

All lanes:

Western blot - Anti-Pannexin 1 antibody [EPR28631-135] (<a href='/en-us/products/primary-antibodies/pannexin-1-antibody-epr28631-135-ab325529'>ab325529</a>) at 1/1000 dilution

Lane 1:

Neuro-2a (mouse neuroblastoma neuroblast) whole cell lysate at 20 µg

Lane 2:

CT26.WT (mouse colon fibroblast) whole cell lysate at 20 µg

Lane 3:

PANC-1 (human pancreatic epithelioid carcinoma epithelial cell) whole cell lysate at 20 µg

Lane 4:

U-937 (human histiocytic lymphoma monocyte) whole cell 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: 37-50 kDa,124 kDa

false

• WB

Supplier Data

Western blot - Anti-Vinculin antibody [EPR8185] - Loading Control (AB129002)

Low expression : SH-SY5Y (PMID : 25883214).

In Western blot, Anti-Vinculin antibody [EPR8185] (ab129002) staining at 1/10000 dilution.

Blocking and diluting buffer and concentration : 5% NFDM/TBST.

All lanes:

Western blot - Anti-Heparan Sulfate Proteoglycan 2/Perlecan antibody [RM2059] (<a href='/en-us/products/primary-antibodies/heparan-sulfate-proteoglycan-2-perlecan-antibody-rm2059-ab318284'>ab318284</a>) at 1/1000 dilution

Lane 1:

HUVEC (human umbilical vein endothelial cell) whole cell lysate at 20 µg

Lane 2:

SH-SY5Y (human neuroblastoma epithelial cell) whole cell lysate at 20 µg

Lane 3:

EA.hy926 (human somatic cell hybrid endothelial cell) whole cell 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: 469 kDa,124 kDa

false

Exposure time: 37s

• WB

Supplier Data

Western blot - Anti-Vinculin antibody [EPR8185] - Loading Control (AB129002)

Negative control : K-562, Hela (PMID : 11855812).

The expression profile/ molecular weight observed is consistent with what has been described in the literature (PMID : 11855812, 16737958).

In Western blot, Anti-Vinculin antibody [EPR8185] (ab129002) staining at 1/10000 dilution.

All lanes:

Western blot - Anti-MUC17 antibody [EPR28482-85] (<a href='/en-us/products/primary-antibodies/muc17-antibody-epr28482-85-ab316971'>ab316971</a>) at 1/1000 dilution

Lane 1:

AsPC-1 (human Pancreas epithelial cell) whole cell lysate at 20 µg

Lane 2:

K-562 (human chronic myelogenous leukemia lymphoblast) whole cell lysate at 20 µg

Lane 3:

HeLa (human cervical adenocarcinoma epithelial cell) whole cell 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: 500 kDa,124 kDa

false

Exposure time: 180s

• WB

Supplier Data

Western blot - Anti-Vinculin antibody [EPR8185] - Loading Control (AB129002)

The lane 4 was developed using a high sensitivity ECL substrate. The high-sensitivity ECL substrate allows for the detection of proteins in the mid-femtogram range.

In Western blot, Anti-Vinculin antibody [EPR8185] (ab129002) staining at 1/10000 dilution.

Exposure time : Lanes 1-3 : 92 seconds, lane 4 : 180 seconds

All lanes:

Western blot - Anti-CEP290 antibody [EPR28924-76] (<a href='/en-us/products/primary-antibodies/cep290-antibody-epr28924-76-ab318278'>ab318278</a>) at 1/1000 dilution

Lane 1:

HeLa (human cervical adenocarcinoma epithelial cell) transfected with scrambled siRNA control whole cell lysate at 20 µg

Lane 2:

HeLa transfected with siRNA specifically targeting CEP290 whole cell lysate at 20 µg

Lane 3:

K-562 (human chronic myelogenous leukemia lymphoblast) whole cell lysate at 40 µg

Lane 4:

Hepa1-6 (mouse hepatoma epithelial cell) whole cell lysate at 40 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 1/100000 dilution

Observed band size: 290 kDa,124 kDa

false

• WB

Supplier Data

Western blot - Anti-Vinculin antibody [EPR8185] - Loading Control (AB129002)

Samples are non-boiled as boiling may cause protein aggregation.

In Western blot, Anti-Vinculin antibody [EPR8185] (ab129002) staining at 1/10000 dilution.

All lanes:

Western blot - Anti-PIEZO2 antibody [EPR27152-48] (<a href='/en-us/products/primary-antibodies/piezo2-antibody-epr27152-48-ab318187'>ab318187</a>) at 1/1000 dilution

Lane 1:

HeLa (human cervical adenocarcinoma epithelial cell) transfected with scrambled siRNA control whole cell lysate at 20 µg

Lane 2:

HeLa transfected with siRNA specifically targeting PIEZO2 whole cell 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: 320 kDa,124 kDa

false

Exposure time: 92s

• WB

Lab

Western blot - Anti-Vinculin antibody [EPR8185] - Loading Control (AB129002)

Blocking and diluting buffer and concentration : 5% NFDM/TBST.

To minimize protein degradation, the fresh lysates were lysed immediately after harvest and then applied to a gel and transfer membrane for Western blotting as soon as possible.

In Western blot, Anti-Vinculin antibody [EPR8185] (ab129002) (1 : 10000) (124KDa).

All lanes:

Western blot - Anti-ATR antibody [EPR30640-130] (<a href='/en-us/products/primary-antibodies/atr-antibody-epr30640-130-ab325605'>ab325605</a>) at 1/1000 dilution

Lane 1:

HeLa (human cervical adenocarcinoma epithelial cell) whole cell lysate at 20 µg

Lane 2:

293T (human embryonic kidney epithelial cell) whole cell lysate at 20 µg

Lane 3:

Jurkat (human T cell leukemia T lymphocyte from peripheral blood) whole cell lysate at 20 µg

Lane 4:

K-562 (human chronic myelogenous leukemia lymphoblast) whole cell lysate at 48 µg

Lane 5:

MIA PaCa-2 (human pancreas epithelial cell) whole cell lysate at 48 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: 270 kDa,124 kDa

false

Exposure time: 180s

• WB

Supplier Data

Western blot - Anti-Vinculin antibody [EPR8185] - Loading Control (AB129002)

SON can undergo degradation easily.

The expression profile/ molecular weight observed is consistent with what has been described in the literature (PMID : 18952841, 27545680).

In Western blot, Anti-Vinculin antibody [EPR8185] (ab129002) staining at 1/10000 dilution.

All lanes:

Western blot - Anti-SON/DBP5 antibody [EPR28617-69] (<a href='/en-us/products/primary-antibodies/son-dbp5-antibody-epr28617-69-ab317311'>ab317311</a>) at 1/1000 dilution

Lane 1:

HeLa (human cervix adenocarcinoma epithelial cell) transfected with scrambled siRNA control whole cell lysate at 40 µg

Lane 2:

HeLa transfected with siRNA specifically targeting SON whole cell lysate at 40 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 1/100000 dilution

Observed band size: 264 kDa,124 kDa

false

Exposure time: 26s

• WB

Supplier Data

Western blot - Anti-Vinculin antibody [EPR8185] - Loading Control (AB129002)

In lanes 1-3, the lysates were stored at -80°C prior to Western Blotting. The bands beneath the target band (205 kDa) are likey to be degradation products. In lanes 4-6, the lysates were freshly made and used for Western Blotting immediately to minimize protein degradation.

In Western blot, Anti-Vinculin antibody [EPR8185] (ab129002) staining at 1/10000 dilution.

All lanes:

Western blot - Anti-CDK12 antibody [EPR29009-30] (<a href='/en-us/products/primary-antibodies/cdk12-antibody-epr29009-30-ab317746'>ab317746</a>) at 1/1000 dilution

Lane 1:

HeLa (human cervical adenocarcinoma epithelial cell) whole cell lysate at 20 µg

Lane 2:

293T (human embryonic kidney epithelial cell) whole cell lysate at 20 µg

Lane 3:

K-562 (human chronic myelogenous leukemia lymphoblast) whole cell lysate at 20 µg

Lane 4:

HeLa (human cervical adenocarcinoma epithelial cell) whole fresh cell lysate at 20 µg

Lane 5:

293T (human embryonic kidney epithelial cell) whole fresh cell lysate at 20 µg

Lane 6:

K-562 (human chronic myelogenous leukemia lymphoblast) whole fresh cell 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 1/100000 dilution

Observed band size: 205 kDa,124 kDa

false

Exposure time: 180s

• WB

Supplier Data

Western blot - Anti-Vinculin antibody [EPR8185] - Loading Control (AB129002)

In Western blot, Anti-Vinculin antibody [EPR8185] (ab129002) staining at 1/10000 dilution.

All lanes:

Western blot - Anti-CDK12 antibody [EPR29009-30] (<a href='/en-us/products/primary-antibodies/cdk12-antibody-epr29009-30-ab317746'>ab317746</a>) at 1/1000 dilution

Lane 1:

293T (human embryonic kidney epithelial cell) transfected with scrambled siRNA control whole cell lysate at 20 µg

Lane 2:

293T transfected with siRNA specifically targeting CDK12 whole cell 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 1/100000 dilution

Observed band size: 205 kDa,124 kDa

false

Exposure time: 180s

• WB

Supplier Data

Western blot - Anti-Vinculin antibody [EPR8185] - Loading Control (AB129002)

In Western blot, ab316279 was shown to bind specifically to WNK1. Target of interest was observed at 251 kDa in wild-type HAP1 cell lysates (lane 1) with no signal observed at this size in WNK1 knockout cell line (lane 2).

The blot of Lanes 1-3 was developed using a high sensitivity ECL substrate.

In Western blot, Anti-Vinculin antibody [EPR8185] (ab129002) staining at 1/10000 dilution.

Exposure time : Lanes 1-3 : 125 seconds; Lane 4 : 26 seconds.

All lanes:

Western blot - Anti-WNK1 antibody [EPR28125-60] (<a href='/en-us/products/primary-antibodies/wnk1-antibody-epr28125-60-ab316279'>ab316279</a>) at 1/1000 dilution

Lane 1:

Wild-type HAP1 (human chronic myelogenous leukemia near-haploid cell) whole cell lysate at 20 µg

Lane 2:

WNK1 knockout HAP1 whole cell lysate at 20 µg

Lane 3:

293T (human embryonic kidney epithelial cell) whole cell lysate at 20 µg

Lane 4:

HeLa (human cervical adenocarcinoma epithelial cell) whole cell 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: 251 kDa,124 kDa

false

• WB

Supplier Data

Western blot - Anti-Vinculin antibody [EPR8185] - Loading Control (AB129002)

Negative control : kidney (PMID : 11855812).

In Western blot, Anti-Vinculin antibody [EPR8185] (ab129002) staining at 1/10000 dilution.

All lanes:

Western blot - Anti-MUC17 antibody [EPR28482-85] (<a href='/en-us/products/primary-antibodies/muc17-antibody-epr28482-85-ab316971'>ab316971</a>) at 1/1000 dilution

Lane 1:

Human small intestine tissue lysate at 20 µg

Lane 2:

Human kidney 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 1/2000 dilution

Observed band size: 500 kDa,124 kDa

true

Exposure time: 180s

• WB

Supplier Data

Western blot - Anti-Vinculin antibody [EPR8185] - Loading Control (AB129002)

The high-sensitivity ECL substrate allows for the detection of proteins in the mid-femtogram range.

In Western blot, Anti-Vinculin antibody [EPR8185] (ab129002) staining at 1/10000 dilution.

All lanes:

Western blot - Anti-CEP290 antibody [EPR28924-76] (<a href='/en-us/products/primary-antibodies/cep290-antibody-epr28924-76-ab318278'>ab318278</a>) at 1/1000 dilution

Lane 1:

Human placenta tissue lysate at 50 µg

Lane 2:

Human adrenal gland tissue lysate at 50 µg

Secondary

All lanes:

Goat Anti-Rabbit IgG (HRP) with minimal cross-reactivity with human IgG at 1/2000 dilution

Observed band size: 290 kDa,124 kDa

true

Exposure time: 180s