Anti-Wnt3a antibody [EPR21889]

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Wnt3a antibody [EPR21889] (AB219412)

Immunohistochemical analysis of paraffin-embedded rat E14.5 tissue labeling Wnt3a with ab219412 at 1/500 dilution, followed by a ready to use Goat Anti-Rabbit IgG H&L (HRP). Cytoplasmic staining on the hem remnant of rat E14.5 (PMID : 15509764; PMID : 16410414) is observed. Counter stained with Hematoxylin.
Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is a ready to use Goat Anti-Rabbit IgG H&L (HRP).
Perform heat mediated antigen retrieval using ab93684 (Tris/EDTA buffer, pH 9.0).

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Wnt3a antibody [EPR21889] (AB219412)

Immunohistochemical analysis of paraffin-embedded mouse E14.5 tissue labeling Wnt3a with ab219412 at 1/500 dilution, followed by a ready to use Goat Anti-Rabbit IgG H&L (HRP). Cytoplasmic staining on the hem remnant of mouse E14.5 (PMID : 15509764; PMID : 16410414) is observed. Counter stained with Hematoxylin.
Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is a ready to use Goat Anti-Rabbit IgG H&L (HRP).
Perform heat mediated antigen retrieval using ab93684 (Tris/EDTA buffer, pH 9.0).

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-Wnt3a antibody [EPR21889] (AB219412)

Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized L Wnt-3a (Wnt-3a stably transfected mouse subcutaneous connective tissue areolar and adipose fibroblast) cells labeling Wnt3a with ab219412 at 1/100 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing cytoplasmic staining in L Wnt-3A cell line. The nuclear counter stain is DAPI (blue).
Counterstained with ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594) at a 1/200 dilution (red).
The negative control is the secondary antibody only.

• Flow Cyt (Intra)

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Flow Cytometry (Intracellular) - Anti-Wnt3a antibody [EPR21889] (AB219412)

Intracellular flow cytometric analysis of 4% paraformaldehyde-fixed, 90% methanol permeabilized L Wnt-3a (Wnt-3a stably transfected mouse subcutaneous connective tissue areolar and adipose fibroblast) cell line (right)labeling Wnt3a with ab219412 at 1/500 compared with a Rabbit monoclonal IgG (ab172730) (left). Goat anti rabbit IgG (Alexa Fluor^® 488, ab150077), at 1/2000 dilution was used as the secondary antibody.

Two Wnt3a-expressing populations can be seen labeled in this stably transfected cell line.

• IP

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Immunoprecipitation - Anti-Wnt3a antibody [EPR21889] (AB219412)

Wnt3a was immunoprecipitated from 0.35 mg L Wnt-3a (Wnt-3a stably-transfected mouse subcutaneous connective tissue areolar and adipose fibroblast) whole cell lysate with ab219412 at 1/30 dilution. Western blot was performed from the immunoprecipitate using ab219412 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/5000 dilution.
Lane 1 : L Wnt-3a (Wnt-3a stably-transfected mouse subcutaneous connective tissue areolar and adipose fibroblast) whole cell lysate 10 μg (Input).
Lane 2 : ab219412 IP in L Wnt-3A whole cell lysate.
Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab219412 in L Wnt-3a whole cell lysate.
Blocking/Dilution buffer and concentration : 5% NFDM/TBST.
Exposure time : 10 seconds.

All lanes:

Immunoprecipitation - Anti-Wnt3a antibody [EPR21889] (ab219412)

Predicted band size: 39 kDa

false

• WB

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Western blot - Anti-Wnt3a antibody [EPR21889] (AB219412)

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

All lanes:

Western blot - Anti-Wnt3a antibody [EPR21889] (ab219412) at 1/1000 dilution

Lane 1:

Human fetal brain lysate at 10 µg

Lane 2:

Human fetal skin lysate at 10 µg

Lane 3:

HCT 116 (human colorectal carcinoma epithelial cell) whole cell lysate at 10 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

Predicted band size: 39 kDa

false

Exposure time: 3min

• WB

Supplier Data

Western blot - Anti-Wnt3a antibody [EPR21889] (AB219412)

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

All lanes:

Western blot - Anti-Wnt3a antibody [EPR21889] (ab219412) at 1/20000 dilution

Lane 1:

His-tagged human Wnt3 recombinant protein (aa22-355), 40 ng

Lane 2:

His-tagged human Wnt3a recombinant protein (aa19-352), 20 ng

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

Predicted band size: 39 kDa

false

Exposure time: 3min

• WB

Supplier Data

Western blot - Anti-Wnt3a antibody [EPR21889] (AB219412)

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

Exposure times : Lane 1 : 3 minutes; Lane 2 : 3 seconds.

All lanes:

Western blot - Anti-Wnt3a antibody [EPR21889] (ab219412) at 1/1000 dilution

Lane 1:

L Wnt-3a (Wnt-3a stably transfected mouse subcutaneous connective tissue areolar and adipose fibroblast) supernatant at 20 µL

Lane 2:

L Wnt-3a (Wnt-3a stably transfected mouse subcutaneous connective tissue areolar and adipose fibroblast) 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

Predicted band size: 39 kDa

false