Anti-Dicer antibody [EPR27386-39]

• Flow Cyt (Intra)

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Flow Cytometry (Intracellular) - Anti-Dicer antibody [EPR27386-39] (AB315232)

Flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized HEK-293 (human embryonic kidney epithelial cell) cells labelling Dicer with ab315232 at 1/50 dilution (1 ug, Red) compared with a Rabbit monoclonal IgG (ab172730) (Black) isotype control and an unlabelled control (cells without incubation with primary antibody and secondary antibody) (Blue).

Goat Anti-Rabbit IgG (Alexa Fluor® 488, ab150081) at 1/5000 dilution was used as the secondary antibody.

• IP

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Immunoprecipitation - Anti-Dicer antibody [EPR27386-39] (AB315232)

Dicer was immunoprecipitated from 0.35 mg HEK-293 (human embryonic kidney epithelial cell) whole cell lysate with ab315232 at 1/30 dilution (2ug in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab315232 at 1/1000 dilution. VeriBlot for IP secondary antibody(HRP)(ab131366) was used at 1/5000 dilution.

Lane 1 : HEK-293 (human embryonic kidney epithelial cell) whole cell lysate

Lane 2 : ab315232 IP in HEK-293 (human embryonic kidney epithelial cell) whole cell lysate

Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab315232 in HEK-293 whole cell lysate

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

All lanes:

Immunoprecipitation - Anti-Dicer antibody [EPR27386-39] (ab315232) at 1/30 dilution

All lanes:

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

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

false

Exposure time: 180s

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Dicer antibody [EPR27386-39] (AB315232)

Immunohistochemical analysis of paraffin-embedded Mouse testis tissue labeling Dicer with ab315232 at 1/100 (4.99 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on mouse testis. The section was incubated with ab315232 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND® RX instrument Counterstained with Hematoxylin.

Secondary antibody only control : Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Dicer antibody [EPR27386-39] (AB315232)

Immunohistochemical analysis of paraffin-embedded Rat testis tissue labeling Dicer with ab315232 at 1/100 (4.99 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on rat testis. The section was incubated with ab315232 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND® RX instrument Counterstained with Hematoxylin.

Secondary antibody only control : Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Dicer antibody [EPR27386-39] (AB315232)

Immunohistochemical analysis of paraffin-embedded Mouse cerebrum tissue labeling Dicer with ab315232 at 1/100 (5.03 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on mouse cerebrum. The section was incubated with ab315232 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND® RX instrument Counterstained with Hematoxylin.

Secondary antibody only control : Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins

• WB

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Western blot - Anti-Dicer antibody [EPR27386-39] (AB315232)

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

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

Exposure time : Lanes 1, 3 and 4 : 180 seconds, lane 2 : 26 seconds.

All lanes:

Western blot - Anti-Dicer antibody [EPR27386-39] (ab315232) at 1/1000 dilution

Lane 1:

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

Lane 2:

HepG2 (human hepatocellular carcinoma epithelial cell) whole cell lysate at 20 µg

Lane 3:

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

Lane 4:

HEK-293 (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 1/100000 dilution

Observed band size: 250 kDa

false

• WB

Supplier Data

Western blot - Anti-Dicer antibody [EPR27386-39] (AB315232)

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

In Western blot, ab315232 was shown to bind specifically to Dicer. A band was observed at 250 kDa in wild-type HAP1 cell lysates whereas no signal observed at this size in DICER1 knockout cell line.

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

All lanes:

Western blot - Anti-Dicer antibody [EPR27386-39] (ab315232) at 1/1000 dilution

Lane 1:

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

Lane 2:

DICER1 knockout HAP1 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: 250 kDa,124 kDa

false

Exposure time: 37s

• WB

Supplier Data

Western blot - Anti-Dicer antibody [EPR27386-39] (AB315232)

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

Low expression : skeletal muscle (PMID : 11889553).

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

Exposure time : Lanes 1-2 : 26 seconds, lanes 3-7 : 59 seconds.

All lanes:

Western blot - Anti-Dicer antibody [EPR27386-39] (ab315232) at 1/1000 dilution

Lane 1:

Mouse testis tissue lysate at 20 µg

Lane 2:

Mouse skeletal muscle tissue lysate at 20 µg

Lane 3:

Rat testis tissue lysate at 20 µg

Lane 4:

Rat skeletal muscle tissue lysate at 20 µg

Lane 5:

Human brain tissue lysate at 20 µg

Lane 6:

Human testis tissue lysate at 20 µg

Lane 7:

Human skeletal muscle tissue lysate at 20 µg

Secondary

Lanes 1 - 4:

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

Lanes 5 - 7:

Goat Anti-Rabbit IgG (HRP) with minimal cross-reactivity with human IgG at 1/100000 dilution

Observed band size: 250 kDa,124 kDa

false

• WB

Supplier Data

Western blot - Anti-Dicer antibody [EPR27386-39] (AB315232)

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

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

All lanes:

Western blot - Anti-Dicer antibody [EPR27386-39] (ab315232) at 1/1000 dilution

Lane 1:

Mouse brain tissue lysate at 20 µg

Lane 2:

Mouse liver tissue lysate at 20 µg

Lane 3:

Mouse testis tissue lysate at 20 µg

Lane 4:

Rat brain tissue lysate at 20 µg

Lane 5:

Rat liver tissue lysate at 20 µg

Lane 6:

Rat testis 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 1/100000 dilution

Observed band size: 250 kDa

false

Exposure time: 180s