Anti-Dnmt3a antibody [EPR29184-89]

• IP

Supplier Data

Immunoprecipitation - Anti-Dnmt3a antibody [EPR29184-89] (AB323708)

Dnmt3a was immunoprecipitated from 0.35 mg Wild-type HeLa (human cervical adenocarcinoma epithelial cell) whole cell lysate with ab323708 at 1/30 dilution (2ug in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab323708 at 1000 dilution.

Blocking and dilution buffer and concentration : 5% NFDM/TBST

All lanes:

Immunoprecipitation - Anti-Dnmt3a antibody [EPR29184-89] (ab323708) at 1/1000 dilution

Lane 1:

(Input) Wild-type HeLa (human cervical adenocarcinoma epithelial cell) whole cell lysate at 10 µg

Lane 2:

ab323708 at 1/30 IP in Wild-type HeLa (human cervical adenocarcinoma epithelial cell) whole cell lysate at 10 µg

Lane 3:

Rabbit monoclonal IgG (<a href='/en-us/products/primary-antibodies/rabbit-igg-monoclonal-epr25a-isotype-control-ab172730'>ab172730</a>) instead of ab323708 in parental HeLa whole cell lysate at 10 µg

Lane 4:

(Input) DNMT3A knockout HeLa whole cell lysate at 10 µg

Lane 5:

ab323708 at 1/30 IP in DNMT3A knockout HeLa whole cell lysate at 10 µg

Lane 6:

Rabbit monoclonal IgG (<a href='/en-us/products/primary-antibodies/rabbit-igg-monoclonal-epr25a-isotype-control-ab172730'>ab172730</a>) instead of ab323708 in DNMT3A knockout HeLa whole cell lysate at 10 µg

Secondary

All lanes:

Immunoprecipitation - VeriBlot for IP Detection Reagent (HRP) (<a href='/en-us/products/reagents/veriblot-for-ip-detection-reagent-hrp-ab131366'>ab131366</a>) at 1/5000 dilution

Observed band size: 130 kDa,36 kDa

false

Exposure time: 24s

• IP

Supplier Data

Immunoprecipitation - Anti-Dnmt3a antibody [EPR29184-89] (AB323708)

Dnmt3a was immunoprecipitated from 0.35 mg NIH/3T3 (mouse embryonic fibroblast) whole cell lysate with ab323708 at 1/30 dilution (2ug in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab323708 at 1000 dilution.

Blocking and dilution buffer and concentration : 5% NFDM/TBST

All lanes:

Immunoprecipitation - Anti-Dnmt3a antibody [EPR29184-89] (ab323708) at 1/1000 dilution

Lane 1:

(Input) NIH/3T3 (mouse embryonic fibroblast) whole cell lysate at 10 µg

Lane 2:

ab323708 at 1/30 IP in NIH/3T3 (mouse embryonic fibroblast) whole cell lysate at 10 µg

Lane 3:

Rabbit monoclonal IgG (<a href='/en-us/products/primary-antibodies/rabbit-igg-monoclonal-epr25a-isotype-control-ab172730'>ab172730</a>) instead of ab323708 in parental NIH/3T3 whole cell lysate at 10 µg

Secondary

All lanes:

Immunoprecipitation - VeriBlot for IP Detection Reagent (HRP) (<a href='/en-us/products/reagents/veriblot-for-ip-detection-reagent-hrp-ab131366'>ab131366</a>) at 1/5000 dilution

Observed band size: 130 kDa

false

Exposure time: 102s

• WB

Supplier Data

Western blot - Anti-Dnmt3a antibody [EPR29184-89] (AB323708)

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

In Western blot, ab323708 was shown to bind specifically to Dnmt3a. Target of interest was observed at 130 kDa in wild-type HAP1 cell lysates (lane 1) with no signal observed at this size in DNMT3A knockout cell line (lane 2).

The identity of the bands between 25 kDa and 37 kDa (in lanes 1-3) are unknown.

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

Exposure time : Lanes 1-2, 4-5 : 180 seconds, lane 3 : 48 seconds

All lanes:

Western blot - Anti-Dnmt3a antibody [EPR29184-89] (ab323708) at 1/1000 dilution

Lane 1:

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

Lane 2:

DNMT3A knockout HAP1 whole cell lysate at 20 µg

Lane 3:

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

Lane 4:

P815 (mouse mastocytoma mast Cell) whole cell lysate at 20 µg

Lane 5:

NIH/3T3 (mouse embryonic 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

Observed band size: 130 kDa,36 kDa

false

• WB

Supplier Data

Western blot - Anti-Dnmt3a antibody [EPR29184-89] (AB323708)

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

In Western blot, ab323708 was shown to bind specifically to Dnmt3a. Target of interest was observed at 130 kDa in wild-type HeLa cell lysates (lane 1) with no signal observed at this size in DNMT3A knockout cell line (lane 2 knockout cell line ab261793 / knockout cell lysate ab257128).

The identity of the bands between 25 kDa and 37 kDa (in lanes 1-2) are unknown.

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

All lanes:

Western blot - Anti-Dnmt3a antibody [EPR29184-89] (ab323708) at 1/1000 dilution

Lane 1:

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

Lane 2:

DNMT3A knockout HeLa 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: 130 kDa,36 kDa

false

Exposure time: 48s

• WB

Supplier Data

Western blot - Anti-Dnmt3a antibody [EPR29184-89] (AB323708)

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

The identity of the bands higher than 250 kDa (in lane 1) and lower than 100 kDa (in lane 2) are unknown.

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

Exposure time : Lane 1 : 8 seconds, lane 2 : 37 seconds

All lanes:

Western blot - Anti-Dnmt3a antibody [EPR29184-89] (ab323708) at 1/1000 dilution

Lane 1:

Mouse E17 brain tissue lysate at 20 µg

Lane 2:

Mouse placenta 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: 130 kDa,36 kDa

false

• WB

Supplier Data

Western blot - Anti-Dnmt3a antibody [EPR29184-89] (AB323708)

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

This antibody reacts with DNMT3A isoform 1 and DNMT3A isoform 3 but not DNMT3A isoform 2 and DNMT3B.

In Western blot, Anti-6X His tag® antibody [EPR20547] - ChIP Grade (ab213204) staining at 1/5000 dilution.

All lanes:

Western blot - Anti-Dnmt3a antibody [EPR29184-89] (ab323708) at 1/1000 dilution

Lane 1:

Purified human DNMT3A isoform 1 recombinant protein at 10 ng

Lane 2:

Purified human DNMT3A isoform 2 recombinant protein at 10 ng

Lane 3:

Purified human DNMT3A isoform 3 recombinant protein at 10 ng

Lane 4:

Purified human DNMT3B recombinant protein at 10 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

Observed band size: 130 kDa,30-150 kDa

false

Exposure time: 180s