PA5-77945

antibody from Invitrogen Antibodies

Targeting: DNMT3A

Western blot (Supportive data in Antibodypedia)

Immunocytochemistry (Supportive data in Antibodypedia)

Immunoprecipitation (Data presented on provider website)

Immunohistochemistry (Supportive data in Antibodypedia)

Immunoelectron microscopy (Recommended by provider)

Other assay (Supportive data in Antibodypedia)

Antibody Data

Product number

PA5-77945

Provider

Invitrogen Antibodies

Product name

DNMT3A Polyclonal Antibody

Antibody type

Polyclonal

Antigen

Recombinant full-length protein

Description

Positive Control: 293T, HeLa, HepG2, Neuro2A, PC-12, Rat2, CL1-0 Predicted Reactivity: Bovine (92%) Store product as a concentrated solution. Centrifuge briefly prior to opening the vial.

Reactivity

Human, Mouse, Rat

Host

Rabbit

Isotype

IgG

Vial size

100 µL

Concentration

1.28 mg/mL

Storage

Store at 4°C short term. For long term storage, store at -20°C, avoiding freeze/thaw cycles.

Western blot

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Western blot analysis of DNMT3A in whole cell lysate using 30 µg of protein. Samples were separated with 5% SDS-PAGE and incubated with DNMT3A polyclonal antibody (Product # PA5-77945) using a dilution of 1:2000 followed by HRP-conjugated anti-rabbit IgG.

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Western blot analysis of DNMT3A in non-transfected (-) and transfected (+) 293T cells using 30 µg of protein. Samples were separated with 5% SDS-PAGE and incubated with DNMT3A polyclonal antibody (Product # PA5-77945) using a dilution of 1:500 followed by HRP-conjugated anti-rabbit IgG.

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Western blot analysis of DNMT3A in whole cell lysate using 30 µg of protein. Samples were separated with 7.5% SDS-PAGE and incubated with DNMT3A polyclonal antibody (Product # PA5-77945) using a dilution of 1:1000 followed by HRP-conjugated anti-rabbit IgG.

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Western blot analysis of DNMT3A in whole cell lysate using 30 µg of protein. Samples were separated with 7.5% SDS-PAGE and incubated with DNMT3A polyclonal antibody (Product # PA5-77945) using a dilution of 1:1000 followed by HRP-conjugated anti-rabbit IgG.

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Western Blot using DNMT3A Polyclonal Antibody (Product # PA5-77945). Various whole cell extracts (30 µg) were separated by 7.5% SDS-PAGE, and the membrane was blotted with DNMT3A Polyclonal Antibody (Product # PA5-77945) diluted at 1:1,000. The HRP-conjugated anti-rabbit IgG antibody was used to detect the primary antibody.

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Western Blot using DNMT3A Polyclonal Antibody (Product # PA5-77945). Whole cell extract (30 µg) was separated by 7.5% SDS-PAGE, and the membrane was blotted with DNMT3A Polyclonal Antibody (Product # PA5-77945) diluted at 1:1,000. The HRP-conjugated anti-rabbit IgG antibody was used to detect the primary antibody.

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Western Blot using DNMT3A Polyclonal Antibody (Product # PA5-77945). Various whole cell extracts (30 µg) were separated by 5% SDS-PAGE, and the membrane was blotted with DNMT3A Polyclonal Antibody (Product # PA5-77945) diluted at 1:2,000. The HRP-conjugated anti-rabbit IgG antibody was used to detect the primary antibody.

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Knockdown of DNMT3A was achieved by transfecting NTERA-2 cl.D1 with DNMT3A specific siRNAs (Silencer® select Product # s200426, s502236). Western blot analysis (Fig. a) was performed using modified whole cell extracts (1% SDS) from the DNMT3A knockdown cells (lane 3), non-specific scrambled siRNA transfected cells (lane 2) and untransfected cells (lane 1). The blot was probed with DNMT3A Polyclonal Antibody (Product # PA5-77945, 1:2000 dilution) and Goat anti-Rabbit IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A27036, 0.25 µg/mL, 1:4000 dilution). Densitometric analysis of this western blot is shown in histogram (Fig. b). Decrease in signal upon siRNA mediated knock down confirms that antibody is specific to DNMT3A.

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Western blot was performed using Anti-DNMT3A Polyclonal Antibody, (Product # PA5-77945) and a 110 kDa band corresponding to DNMT3A was observed in the cell lines, with higher expression in NTERA-2 compared to HeLa and Jurkat. Modified whole cell extracts (1% SDS) (40 µg lysate) of NTERA-2 (Lane 1), HeLa (Lane 2), MCF7 (Lane 3), Jurkat (Lane 4), NCCIT (lane 5) and iPSC (Lane 6) were electrophoresed using Novex® NuPAGE® 4-12 % Bis-Tris gel (Product # NP0321BOX). Resolved proteins were then transferred onto a nitrocellulose membrane (Product # IB23001) by iBlot® 2 Dry Blotting System (Product # IB21001). The blot was probed with the primary antibody (1:2000 dilution) and detected by chemiluminescence with Goat anti-Rabbit IgG (H+L), Superclonal™ Recombinant Secondary Antibody, HRP (Product # A27036, 1:4000 dilution) using the iBright FL 1000 (Product # A32752). Chemiluminescent detection was performed using Novex® ECL Chemiluminescent Substrate Reagent Kit (Product # WP20005).

Immunocytochemistry

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Immunocytochemistry-Immunofluorescence analysis of DNMT3A was performed in 293T cells fixed in 4% paraformaldehyde at RT for 15 min. Green: DNMT3A Polyclonal Antibody (Product # PA5-77945) diluted at 1:1000. Red: phalloidin, a cytoskeleton marker. Scale bar = 10 µm.

Immunohistochemistry

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

DNMT3A Polyclonal Antibody detects DNMT3A protein at nucleus on mouse ovary by immunohistochemical analysis. Sample: Paraffin-embedded mouse ovary. DNMT3A Polyclonal Antibody (Product # PA5-77945) dilution: 1:500. Antigen Retrieval: EDTA based buffer, pH 8.0, 15 min.

Other assay

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

DNMT3A Polyclonal Antibody immunoprecipitates DNMT3A protein in IP experiments. IP samples: 293T nuclear extract. A. 50 µg 293T nuclear extract. B. Control with 4 µg of preimmune Rabbit IgG. C. Immunoprecipitation of DNMT3A protein by 4 µg DNMT3A Polyclonal Antibody (Product # PA5-77945). 5 % SDS-PAGE. The immunoprecipitated DNMT3A protein was detected by DNMT3A Polyclonal Antibody (Product # PA5-77945) diluted at 1:2,000.

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Immunoprecipitation of DNMT3A was performed in 293T whole cell extracts using 5 µg of DNMT3A Polyclonal Antibody (Product # PA5-77945). Samples were transferred to a membrane and probed with DNMT3A Polyclonal Antibody as a primary antibody and an HRP-conjugated anti-Rabbit IgG was used as a secondary antibody.