重组Anti-alpha Actinin 4抗体[EPR2533(2)]

• Flow Cyt (Intra)

Supplier Data

Flow Cytometry (Intracellular) - Anti-alpha Actinin 4 antibody [EPR2533(2)] - Loading Control (AB108198)

Flow Cytometry analysis of HeLa (Human cervix adenocarcinoma epithelial cell) cells labelling alpha Actinin 4 with Purified ab108198 at 1 : 50 dilution (10 µg/ml) (Red). Cells were fixed with 4% Paraformaldehyde and permeabilised with 90% Methanol. A Goat anti rabbit IgG (Alexa Fluor® 488, ab150077) secondary antibody was used at 1 : 2000. Isotype control - Rabbit monoclonal IgG (Black). Unlabelled control - Cell without incubation with primary antibody and secondary antibody (Blue).

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-alpha Actinin 4 antibody [EPR2533(2)] - Loading Control (AB108198)

ab108198 staining ACTN4 in wild-type HAP1 cells (top panel) and ACTN4 knockout HAP1 cells (bottom panel). The cells were fixed with PFA (10min), permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated with ab108198 at 1/100 dilution and ab7291 (Tubulin) at 1/1000 dilution overnight at +4°C, followed by a further incubation at room temperature for 1h with a goat secondary antibody to rabbit IgG (Alexa Fluor® 488) (ab150081) at 2 μg/ml (shown in green) and a goat secondary antibody to mouse IgG (Alexa Fluor® 594) (ab150120) at 2 μg/ml (shown in pseudo color red). Nuclear DNA was labelled in blue with DAPI.

Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-alpha Actinin 4 antibody [EPR2533(2)] - Loading Control (AB108198)

Immunocytochemistry analysis of MCF7 (Human breast adenocarcinoma epithelial cell) cells labeling alpha Actinin 4 with Purified ab108198 at 1 : 250 dilution (2.0 µg/ml). Cells were fixed in 4% Paraformaldehyde and permeabilized with 0.1% tritonX-100. Cells were counterstained with ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594) 1 : 200 (2.5 µg/ml). Goat anti rabbit IgG (Alexa Fluor® 488, ab150077) was used as the secondary antibody at 1 : 1000 (2 µg/ml) dilution. DAPI (blue) was used as nuclear counterstain. PBS instead of the primary antibody was used as the secondary antibody only control.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-alpha Actinin 4 antibody [EPR2533(2)] - Loading Control (AB108198)

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human breast carcinoma tissue sections labeling alpha Actinin 4 with Purified ab108198 at 1 : 150 dilution (3.29 µg/mL). Heat mediated antigen retrieval was performed using Perform heat mediated antigen retrieval using ab93684 (Tris/EDTA buffer, pH 9.0). Tissue was counterstained with Hematoxylin. ImmunoHistoProbe one step HRP Polymer (ready to use) secondary antibody was used. PBS instead of the primary antibody was used as the negative control.

• IP

Unknown

Immunoprecipitation - Anti-alpha Actinin 4 antibody [EPR2533(2)] - Loading Control (AB108198)

Purified ab108198 at 1/50 dilution (2μg) immunoprecipitating alpha Actinin 4 in HeLa whole cell lysate.
Lane 1 (input) : HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysate 10μg
Lane 2 (+) : ab108198 + HeLa whole cell lysate.
Lane 3 (-) : Rabbit monoclonal IgG (ab172730) instead of ab108198 in HeLa whole cell lysate.
VeriBlot for IP Detection Reagent (HRP) (ab131366) (1/1000 dilution) was used for Western blotting.
Blocking Buffer and concentration : 5% NFDM/TBST.
Diluting buffer and concentration : 5% NFDM/TBST.
Observed band size : 105 kDa

All lanes:

Immunoprecipitation - Anti-alpha Actinin 4 antibody [EPR2533(2)] - Loading Control (ab108198)

Predicted band size: 105 kDa

false

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-alpha Actinin 4 antibody [EPR2533(2)] - Loading Control (AB108198)

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of mouse liver tissue sections labeling alpha Actinin 4 with Purified ab108198 at 1 : 150 dilution (3.29 µg/mL). Heat mediated antigen retrieval was performed using Perform heat mediated antigen retrieval using ab93684 (Tris/EDTA buffer, pH 9.0). Tissue was counterstained with Hematoxylin. ImmunoHistoProbe one step HRP Polymer (ready to use) secondary antibody was used. PBS instead of the primary antibody was used as the negative control.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-alpha Actinin 4 antibody [EPR2533(2)] - Loading Control (AB108198)

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of rat liver tissue sections labeling alpha Actinin 4 with Purified ab108198 at 1 : 150 dilution (3.29 µg/mL). Heat mediated antigen retrieval was performed using Perform heat mediated antigen retrieval using ab93684 (Tris/EDTA buffer, pH 9.0). Tissue was counterstained with Hematoxylin. ImmunoHistoProbe one step HRP Polymer (ready to use) secondary antibody was used. PBS instead of the primary antibody was used as the negative control.

• WB

Supplier Data

Western blot - Anti-alpha Actinin 4 antibody [EPR2533(2)] - Loading Control (AB108198)

Negative control : 293T (PMID : 20444890). The expression profile is consistent with what has been described in the literature (PMID : 20444890). This blot was produced using a 4-12% Bis-Tris gel and MOPS buffer. The gel was run at 200V for 54 minutes and then transferred to a Nitrocellulose membrane at 25V for 10 minutes. The membrane was blocked for an hour using 3% milk and incubated overnight at 4°C with Mouse monoclonal [CM2B4] to Polyoma virus, Large T antigen ( ab307450) and Rabbit anti-ACTN4 (loading control, ab108198) at a 1/1000 and 1/20 000 dilution, respectively. Antibody binding was detected using Goat Anti-Mouse IgG H&L (IRDye® 800CW, green) and Goat Anti-Rabbit IgG H&L (IRDye® 680RD, red) secondary antibodies at a 1/20 000 dilution (1h at room temperature) before imaging. Blocking/Dilution buffer : 3% NFDM/TBST.

All lanes:

Western blot - Anti-Polyoma virus, Large T antigen antibody [CM2B4] (<a href='/products/primary-antibodies/polyoma-virus-large-t-antigen-antibody-cm2b4-ab307450'>ab307450</a>) at 1/1000 dilution

Lane 1:

MKL-1 whole cell lysate at 20 µg

Lane 2:

293T whole cell lysate at 20 µg

Secondary

All lanes:

Western blot - Goat anti-Mouse IgG H&L (IRDye® 800CW) preadsorbed (<a href='/products/secondary-antibodies/goat-mouse-igg-h-l-irdye-800cw-preadsorbed-ab216772'>ab216772</a>) at 1/20000 dilution

Observed band size: 48 kDa

false

• WB

Lab

Western blot - Anti-alpha Actinin 4 antibody [EPR2533(2)] - Loading Control (AB108198)

Western blot : Mouse Monoclonal[AMACR/1864] to AMACR ab268062 staining at 2 µg/mL, shown in green; Rabbit anti-ACTN4 (ab108198) loading control staining at 1/20,000 dilution, shown in magenta. A band was observed at 40 kDa in Wild-type U-87 MG cell lysates with no signal observed at this size in AMACR knockout U-87 MG cell line. To generate this image, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3pc 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-Mouse 800CW & Goat anti-Rabbit 680RD at 1/20,000 dilution.

All lanes:

Western blot - Anti-AMACR antibody [AMACR/1864] (<a href='/products/primary-antibodies/amacr-antibody-amacr-1864-ab268062'>ab268062</a>) at 2 µg/mL

Lane 1:

Wild-type U-87 MG at 20 µg

Lane 2:

AMACR knockout U-87 MG at 20 µg

Lane 3:

LNCaP at 20 µg

Lane 4:

Raji at 20 µg

Secondary

Lanes 1 - 4:

Goat anti-Mouse 800CW at 1/20000 dilution

Lanes 1 - 4:

Goat anti-Rabbit 680RD at 1/20000 dilution

Predicted band size: 42 kDa

Observed band size: 40 kDa,100 kDa

false

• WB

Lab

Western blot - Anti-alpha Actinin 4 antibody [EPR2533(2)] - Loading Control (AB108198)

Anti-CD38 antibody [5C5C3] - Extracellular domain staining at 1/500 dilution, shown in green; Rabbit anti-ACTN4 [EPR2533(2)] (ab108198) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab204940 was shown to bind specifically to CD38. A band was observed at 42 kDa in wild-type A549 cell lysates with no signal observed at this size in CD38 knockout cell line. To generate this image, wild-type and CD38 knockout A549 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-Mouse IgG H&L 800CW and Goat anti-Rabbit IgG H&L 680RD at 1/20000 dilution.

All lanes:

Western blot - Anti-CD38 antibody [5C5C3] - Extracellular domain (<a href='/products/primary-antibodies/cd38-antibody-5c5c3-extracellular-domain-ab204940'>ab204940</a>) at 1/500 dilution

Lane 1:

Wild-type A549 cell lysate at 20 µg

Lane 2:

Western blot - Human CD38 knockout A549 cell line (<a href='/products/cell-lines/human-cd38-knockout-a549-cell-line-ab289025'>ab289025</a>)

Lane 2:

CD38 knockout A549 cell lysate at 20 µg

Lane 3:

Daudi cell lysate at 20 µg

Lane 4:

HCT 116 cell lysate at 20 µg

Secondary

All lanes:

Goat anti-Mouse IgG H&L 800CW and Goat anti-Rabbit IgG H&L 680RD at 1/20000 dilution

Predicted band size: 34 kDa

Observed band size: 42 kDa

false

• WB

Lab

Western blot - Anti-alpha Actinin 4 antibody [EPR2533(2)] - Loading Control (AB108198)

Western blot : Mouse Monoclonal[10/Fibronectin] to Fibronectin ab281575 staining at 1/500 dilution, shown in green; Rabbit anti-ACTN4 (ab108198) loading control staining at 1/20,000 dilution, shown in magenta.

A band was observed at 272 kDa in Wild-type A549 cell lysates with no signal observed at this size in FN1 knockout A549 cell line.

To generate this image, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3pc 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 800CW & Goat anti-Mouse 680RD at 1/20,000 dilution.

All lanes:

Western blot - Anti-Fibronectin antibody [10/Fibronectin] (<a href='/products/primary-antibodies/fibronectin-antibody-10-fibronectin-ab281575'>ab281575</a>) at 1/500 dilution

Lane 1:

Wild-type A549 at 20 µg

Lane 2:

FN1 knockout A549

Lane 2:

Western blot - Human FN1 knockout A549 cell line (<a href='/products/cell-lines/human-fn1-knockout-a549-cell-line-ab286514'>ab286514</a>) at 20 µg

Lane 3:

HepG2 at 20 µg

Secondary

All lanes:

Goat anti-Rabbit 800CW & Goat anti-Mouse 680RD at 1/20000 dilution

Predicted band size: 272 kDa

Observed band size: 272 kDa

false

• WB

Supplier Data

Western blot - Anti-alpha Actinin 4 antibody [EPR2533(2)] - Loading Control (AB108198)

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 : 16721827).

In Western blot, Anti-alpha Actinin 4 antibody [EPR2533 (2)] - Loading Control (ab108198) staining at 1/200000 dilution (104 kDa).

All lanes:

Western blot - Anti-CDK13 antibody [EPR29353-534] (<a href='/products/primary-antibodies/cdk13-antibody-epr29353-534-ab323865'>ab323865</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 CDK13 whole cell 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: 200 kDa,104 kDa

false

Exposure time: 10s

• WB

Supplier Data

Western blot - Anti-alpha Actinin 4 antibody [EPR2533(2)] - Loading Control (AB108198)

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

Negative control : THP-1; MOLT-4

The molecular weight observed is consistent with what has been described in the literature (PMID : 33179593; PMID : 33179593).

In Western blot, Anti-alpha Actinin 4 antibody [EPR2533 (2)](ab108198) staining at 105KDa dilution.

All lanes:

Western blot - Anti-KIF13A antibody [EPR29130-81] (<a href='/products/primary-antibodies/kif13a-antibody-epr29130-81-ab323873'>ab323873</a>) at 1/1000 dilution

Lane 1:

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

Lane 2:

TF-1 (human erythroleukemia erythroblast) whole cell lysate at 40 µg

Lane 3:

THP-1 (human monocytic leukemia monocyte) whole cell lysate at 40 µg

Lane 4:

MOLT-4 (human lymphoblastic leukemia T lymphoblast) whole cell lysate at 40 µg

Secondary

All lanes:

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

Observed band size: 250 kDa,105 kDa

false

Exposure time: 180s

• WB

Supplier Data

Western blot - Anti-alpha Actinin 4 antibody [EPR2533(2)] - Loading Control (AB108198)

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

Negative control : Daudi, Jurkat, U-937, HL-60.

The molecular weight observed is consistent with what has been described in the literature (PMID : 33179593; PMID : 33179593).

In Western blot, Anti-alpha Actinin 4 antibody [EPR2533 (2)](ab108198) staining at 105KDa dilution.

All lanes:

Western blot - Anti-KIF13A antibody [EPR29130-81] (<a href='/products/primary-antibodies/kif13a-antibody-epr29130-81-ab323873'>ab323873</a>) at 1/1000 dilution

Lane 1:

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

Lane 2:

LN-229 (human brain glioblastoma epithelial cell) whole cell lysate at 40 µg

Lane 3:

Daudi (human Burkitt's lymphoma lymphoblast) whole cell lysate at 40 µg

Lane 4:

U-937 (human histiocytic lymphoma monocyte) whole cell lysate at 40 µg

Lane 5:

Jurkat (human T cell leukemia T lymphocyte from peripheral blood) whole cell lysate at 40 µg

Lane 6:

HL-60 (human acute promyelocytic leukemia promyeloblast) whole cell lysate at 40 µg

Secondary

All lanes:

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

Observed band size: 250 kDa,105 kDa

false

Exposure time: 180s

• WB

Lab

Western blot - Anti-alpha Actinin 4 antibody [EPR2533(2)] - Loading Control (AB108198)

Western blot : Anti-RAS antibody [18/Ras] ab307738 staining at 1/200 dilution, shown in green; Rabbit anti-ACTN4 ab108198 loading control staining at 1/20000 dilution, shown in magenta. A band was observed at 22 kDa in Wild-type MCF7 cell lysates with no signal observed at this size in HRAS knockout MCF7 cell line. To generate this image, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3pc 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-Mouse 800CW and Goat anti-Rabbit 680RD at 1/20,000 dilution.

All lanes:

Western blot - Anti-RAS antibody [18/Ras] (<a href='/products/primary-antibodies/ras-antibody-18-ras-ab307738'>ab307738</a>) at 1/200 dilution

Lane 1:

Wild-type MCF7 whole cell lysate at 20 µg

Lane 2:

HRAS knockout MCF7 whole cell lysate at 20 µg

Lane 3:

HEK-293 whole cell lysate at 20 µg

Lane 4:

HeLa whole cell lysate at 20 µg

Secondary

All lanes:

Goat anti-Mouse 800CW & Goat anti-Rabbit 680RD at 1/20000 dilution

Predicted band size: 21 kDa

Observed band size: 22 kDa

false

• WB

Lab

Western blot - Anti-alpha Actinin 4 antibody [EPR2533(2)] - Loading Control (AB108198)

Western blot : Anti-LIPA antibody [1F9] (ab219113) staining at 1/1000 dilution, shown in green; Rabbit anti-ACTN4 [EPR2533(2)] (ab108198) loading control staining at 1/20000 dilution, shown in magenta. In Western blot, ab219113 was shown to bind specifically to LIPA. A band was observed at 54 kDa in wild-type A549 cell lysates with no signal observed at this size in LIPA knockout cell line. To generate this image, wild-type and LIPA knockout A549 cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 5 % 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-Mouse IgG H&L 800CW and Goat anti-Rabbit IgG H&L 680RD at 1/20000 dilution.

All lanes:

Western blot - Anti-Lysosomal acid lipase/LAL antibody [1F9] (<a href='/products/primary-antibodies/lysosomal-acid-lipase-lal-antibody-1f9-ab219113'>ab219113</a>) at 1/1000 dilution

Lane 1:

Wild-type A549 cell lysate at 20 µg

Lane 2:

LIPA knockout A549 cell lysate at 20 µg

Secondary

Lanes 1 - 2:

Goat anti-Mouse IgG H&L 800CW at 1/20000 dilution

Lanes 1 - 2:

Goat anti-Rabbit IgG H&L 680RD at 1/20000 dilution

Observed band size: 54 kDa

false

• WB

Lab

Western blot - Anti-alpha Actinin 4 antibody [EPR2533(2)] - Loading Control (AB108198)

Western blot : Mouse Monoclonal[10/Fibronectin] to Fibronectin ab281575 staining at 1/500 dilution, shown in green; Rabbit anti-ACTN4 (ab108198) loading control staining at 1/20,000 dilution, shown in magenta. A band was observed at 238-268 kDa in Wild-type MCF7 cell lysates with no signal observed at this size in FN1 knockout MCF7 cell line. To generate this image, 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-Mouse 800CW & Goat anti-Rabbit 680RD at 1/20,000 dilution.

All lanes:

Western blot - Anti-Fibronectin antibody [10/Fibronectin] (<a href='/products/primary-antibodies/fibronectin-antibody-10-fibronectin-ab281575'>ab281575</a>) at 1/500 dilution

Lane 1:

Wild-type MCF7 at 20 µg

Lane 2:

Western blot - Human FN1 knockout MCF7 cell line (ab286324) at 20 µg

Lane 3:

HepG2 at 20 µg

Lane 4:

PC-3 at 20 µg

Secondary

All lanes:

Goat anti-Mouse 800CW & Goat anti-Rabbit 680RD at 1/20000 dilution

Predicted band size: 272 kDa

Observed band size: 238-268 kDa

false

• WB

Unknown

Western blot - Anti-alpha Actinin 4 antibody [EPR2533(2)] - Loading Control (AB108198)

All lanes:

Western blot - Anti-alpha Actinin 4 antibody [EPR2533(2)] - Loading Control (ab108198) at 1/1000 dilution

Lane 1:

Human skeletal muscle lysate at 20 µg

Lane 2:

A431 (Human epidermoid carcinoma epithelial cell) whole cell lysate at 20 µg

Lane 3:

Mouse brain lysate at 20 µg

Lane 4:

Rat brain lysate at 20 µg

Lane 5:

Rat heart lysate at 20 µg

Secondary

All lanes:

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

Predicted band size: 105 kDa

Observed band size: 105 kDa

false

• WB

Unknown

Western blot - Anti-alpha Actinin 4 antibody [EPR2533(2)] - Loading Control (AB108198)

All lanes:

Western blot - Anti-alpha Actinin 4 antibody [EPR2533(2)] - Loading Control (ab108198) at 1/10000 dilution

All lanes:

HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysate at 15 µg

Secondary

All lanes:

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

Predicted band size: 105 kDa

Observed band size: 105 kDa

false

• WB

Lab

Western blot - Anti-alpha Actinin 4 antibody [EPR2533(2)] - Loading Control (AB108198)

Western blot : Anti-CK1 epsilon antibody [1/Casein Kinase Iepsilon] ab302638 staining at 1/1000 dilution, shown in green; Rabbit anti-ACTN4 ab108198 loading control staining at 1/20000 dilution, shown in magenta. A band was observed at 47 kDa in Wild-type A549 cell lysates with no signal observed at this size in CSNK1E knockout A549 cell line. To generate this image, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 5pc 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-Mouse 800CW and Goat anti-Rabbit 680RD at 1/20,000 dilution.

All lanes:

Western blot - Anti-CK1 epsilon antibody [1/Casein Kinase Iε] (<a href='/products/primary-antibodies/ck1-epsilon-antibody-1-casein-kinase-i-ab302638'>ab302638</a>) at 1/1000 dilution

Lane 1:

Wild-type A549 at 20 µg

Lane 2:

CSNK1E knockout A549 at 20 µg

Lane 3:

Jurkat at 20 µg

Lane 4:

PC-3 at 20 µg

Secondary

All lanes:

Goat anti-Mouse 800CW & Goat anti-Rabbit 680RD at 1/20000 dilution

Predicted band size: 47 kDa

Observed band size: 47 kDa,110 kDa

false

• WB

Lab

Western blot - Anti-alpha Actinin 4 antibody [EPR2533(2)] - Loading Control (AB108198)

Lane 1 : Wild-type HAP1 whole cell lysate (20 μg)
Lane 2 : ACTN4 (alpha Actinin 4) knockout HAP1 whole cell lysate (20 μg)
Lane 3 : HeLa whole cell lysate (20 μg)
Lane 4 : MCF7 whole cell lysate (20 μg)

Lanes 1 - 4 : Merged signal (red and green). Green - ab108198 observed at 105 kDa. Red - loading control, ab9484, observed at 37 kDa.

ab108198 was shown to specifically react with alpha Actinin 4 in wild-type HAP1 cells as signal was lost in ACTN4 (alpha Actinin 4) knockout cells. Wild-type and ACTN4 (alpha Actinin 4) knockout samples were subjected to SDS-PAGE. ab108198 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.

All lanes:

Western blot - Anti-alpha Actinin 4 antibody [EPR2533(2)] - Loading Control (ab108198)

Predicted band size: 105 kDa

false

• WB

Supplier Data

Western blot - Anti-alpha Actinin 4 antibody [EPR2533(2)] - Loading Control (AB108198)

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

Negative : HEL, 786-O.

PTCH1 is a glycosylated protein and can be deglycosylated by Protein Deglycosylation MIX II.

In Western blot, Anti-alpha Actinin 4 antibody [EPR2533 (2)] - Loading Control (ab108198) staining at 1/100,000 dilution.

All lanes:

Western blot - Anti-Patched / PTCH1 antibody [EPR26072-78] (<a href='/products/primary-antibodies/patched-ptch1-antibody-epr26072-78-ab325234'>ab325234</a>) at 1/1000 dilution

Lane 1:

Untreated 22Rv1 (human prostate carcinoma epithelial cell) whole cell lysate at 20 µg

Lane 2:

22Rv1 whole cell lysate treated with Protein Deglycosylation MIX II at 20 µg

Lane 3:

HEL (human erythroleukemia erythroblast) whole cell lysate at 20 µg

Lane 4:

786-O (human kidney epithelial cell) whole cell lysate at 20 µg

Lane 5:

IMR-32 (human neuroblastoma neuroblast) whole cell lysate at 20 µg

Secondary

All lanes:

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

Observed band size: 160-180 kDa,105 kDa

false

Exposure time: 180s