Anti-DRP1 antibody [EPR19274]

• WB

Supplier Data

Western blot - Anti-DRP1 antibody [EPR19274] (AB184247)

Blocking/Dilution buffer : 5% NFDM/TBST.

Exposure time : Lane 1 and 2 : 3 minutes; Lane 3 : 30 seconds; Lane 4,5 and 6 : 8 seconds.

DRP1 can be SUMOylated, as described in the literature (PMID : 19638400).

All lanes:

Western blot - Anti-DRP1 antibody [EPR19274] (ab184247) at 1/1000 dilution

Lane 1:

A549 (Human lung carcinoma cell line) whole cell lysate at 10 µg

Lane 2:

U-2 OS (Human bone osteosarcoma epithelial cell line) whole cell lysate at 10 µg

Lane 3:

HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate at 10 µg

Lane 4:

Jurkat (Human T cell leukemia cell line from peripheral blood) whole cell lysate at 10 µg

Lane 5:

HEK-293 (Human epithelial cell line from embryonic kidney) whole cell lysate at 10 µg

Lane 6:

HCT 116 (Human colorectal carcinoma cell line) 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: 82 kDa

Observed band size: 83 kDa

false

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-DRP1 antibody [EPR19274] (AB184247)

Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HeLa (Human epithelial cell line from cervix adenocarcinoma) cells labeling DRP1 with ab184247 at 1/250 dilution, followed by Goat anti-Rabbit IgG (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing cytoplasm staining on HeLa cell line. The nuclear counter stain is DAPI (blue).

Tubulin is detected with Anti-alpha Tubulin antibody [EPR19274] - Loading Control (ab7291) at 1/1000 dilution and Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) preadsorbed (ab150120) at 1/1000 dilution (red).

The negative controls are as follows :
-ve control 1 : ab184247 at 1/250 dilution followed by ab150120 at 1/1000 dilution.
-ve control 2 : ab7291 at 1/1000 dilution followed by ab150077 at 1/1000 dilution.

• IP

Supplier Data

Immunoprecipitation - Anti-DRP1 antibody [EPR19274] (AB184247)

DRP1 was immunoprecipitated from 1mg of HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate with ab184247 at 1/30 dilution. Western blot was performed from the immunoprecipitate using ab184247 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/10000 dilution.
Lane 1 : HeLa whole cell lysate 10μg (Input).
Lane 2 : ab184247 IP in HeLa whole cell lysate.
Lane 3 : Rabbit IgG,monoclonal [EPR19274]-Isotype Control (ab172730) instead of ab184247 in HeLa whole cell lysate.
Blocking and dilution buffer and concentration : 5% NFDM/TBST.
Exposure time : 5 seconds.

Note : DRP1 can be SUMOylated, as described in the literature (PMID : 19638400).

All lanes:

Immunoprecipitation - Anti-DRP1 antibody [EPR19274] (ab184247)

Predicted band size: 82 kDa

false

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-DRP1 antibody [EPR19274] (AB184247)

Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized NIH/3T3 (Mouse embryonic fibroblast cell line) cells labeling DRP1 with ab184247 at 1/250 dilution, followed by Goat Anti-Rabbit IgG (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing cytoplasm staining on NIH/3T3 cell line. The nuclear counter stain is DAPI (blue).

Tubulin is detected with Anti-alpha Tubulin antibody [EPR19274] - Loading Control (ab7291) at 1/1000 dilution and Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) preadsorbed (ab150120) at 1/1000 dilution (red).

The negative controls are as follows :
-ve control 1 : ab184247 at 1/250 dilution followed by ab150120 at 1/1000 dilution.
-ve control 2 : ab7291 at 1/1000 dilution followed by ab150077 at 1/1000 dilution.

• Flow Cyt (Intra)

Supplier Data

Flow Cytometry (Intracellular) - Anti-DRP1 antibody [EPR19274] (AB184247)

Intracellular flow cytometric analysis of 4% paraformaldehyde-fixed NIH/3T3 (Mouse embryonic fibroblast cell line) cells labeling DRP1 with ab184247 at 1/70 dilution (red) compared with a Rabbit IgG,monoclonal -Isotype Control (ab172730; black) and an unlabelled control (cells without incubation with primary antibody and secondary antibody; blue). Goat anti rabbit IgG (FITC) at 1/500 dilution was used as the secondary antibody.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-DRP1 antibody [EPR19274] (AB184247)

Immunohistochemical analysis of paraffin-embedded Rat cerebellum tissue labeling DRP1 with ab184247 at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution. Cytoplasm staining on rat cerebellum is observed. Counter stained with Hematoxylin.

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-DRP1 antibody [EPR19274] (AB184247)

Immunohistochemical analysis of paraffin-embedded Mouse cerebrum tissue labeling DRP1 with ab184247 at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution. Cytoplasm staining on mouse cerebrum is observed. Counter stained with Hematoxylin.

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

• WB

Supplier Data

Western blot - Anti-DRP1 antibody [EPR19274] (AB184247)

Blocking/Dilution buffer : 5% NFDM/TBST.

All lanes:

Western blot - Anti-DRP1 antibody [EPR19274] (ab184247) at 1/1000 dilution

All lanes:

Human fetal kidney lysate at 10 µg

Secondary

All lanes:

Goat Anti-Rabbit IgG Peroxidase Conjugate, specific to the non-reduced form of IgG at 1/100000 dilution

Predicted band size: 53 kDa,56 kDa,82 kDa

Observed band size: 53 kDa,56 kDa,83 kDa

false

Exposure time: 30s

• WB

Supplier Data

Western blot - Anti-DRP1 antibody [EPR19274] (AB184247)

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

The identity of the lower MW band at approximately 20 kDa is unknown.

The expression profile/ molecular weight observed is consistent with what has been described in the literature (PMID : 19436752).

ab184247 was used as the loading control at a 1/1000 dilution.

All lanes:

Western blot - Anti-DRP1 (phospho S616) antibody [EPR27387-57] (<a href='/en-us/products/primary-antibodies/drp1-phospho-s616-antibody-epr27387-57-ab314755'>ab314755</a>) at 1/1000 dilution

Lane 1:

Untreated HAP1 (human chronic myelogenous leukemia near-haploid cell line) whole cell lysate (untreated membrane) at 20 µg

Lane 2:

HAP1 treated with 100 ng/ml nocodazole for 16 hours whole cell lysate (untreated membrane) at 20 µg

Lane 3:

Untreated HAP1 (human chronic myelogenous leukemia near-haploid cell line) whole cell lysate (phosphatase treated membrane) at 20 µg

Lane 4:

HAP1 treated with 100 ng/ml nocodazole for 16 hours whole cell lysate (phosphatase treated membrane) 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: 82 kDa

Observed band size: 83 kDa

false

Exposure time: 92s

• WB

Supplier Data

Western blot - Anti-DRP1 antibody [EPR19274] (AB184247)

Blocking and diluting buffer : 5% NFDM/TBST.

ab181602 was used as a loading control at 1/1000000 dilution.

ab184248 worked better in western blot application.

Lanes 1 - 4:

Western blot - Anti-DRP1 antibody [EPR19274] (ab184247) at 1/1000 dilution

Lanes 5 - 8:

Western blot - Anti-DRP1 antibody [EPR19275] (<a href='/en-us/products/primary-antibodies/drp1-antibody-epr19275-ab184248'>ab184248</a>) at 1/1000 dilution

Lanes 1 and 5:

Mouse brain tissue lysate at 20 µg

Lanes 2 and 6:

Mouse liver tissue lysate at 20 µg

Lanes 3 and 7:

Mouse heart tissue lysate at 20 µg

Lanes 4 and 8:

Mouse lung 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

Predicted band size: 82 kDa

Observed band size: 80 kDa

false

Exposure time: 7s

• WB

Supplier Data

Western blot - Anti-DRP1 antibody [EPR19274] (AB184247)

Blocking/Dilution buffer : 5% NFDM/TBST.

Lane 1 : 2 seconds; Lane 2 : 8 seconds; Lane 3 : 3 seconds.

All lanes:

Western blot - Anti-DRP1 antibody [EPR19274] (ab184247) at 1/1000 dilution

Lane 1:

Rat brain lysate at 10 µg

Lane 2:

Rat heart lysate at 10 µg

Lane 3:

Mouse brain 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: 82 kDa

Observed band size: 83 kDa

false

• WB

Supplier Data

Western blot - Anti-DRP1 antibody [EPR19274] (AB184247)

Blocking/Dilution buffer : 5% NFDM/TBST.

All lanes:

Western blot - Anti-DRP1 antibody [EPR19274] (ab184247) at 1/1000 dilution

Lane 1:

PC-12 (Rat adrenal gland pheochromocytoma cell line) whole cell lysate at 10 µg

Lane 2:

NIH/3T3 (Mouse embryonic fibroblast cell line) 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: 141 kDa,42 kDa,82 kDa,86 kDa,94 kDa

Observed band size: 83 kDa,86 kDa

false

Exposure time: 3min

• WB

Supplier Data

Western blot - Anti-DRP1 antibody [EPR19274] (AB184247)

Blocking and diluting buffer and concentration : 5% NFDM/TBST. In western blot, anti-GAPDH antibody [EPR16891] - Loading Control (ab181602) staining at 1/200000 dilution. In western blot, anti-DRP1 antibody [EPR19274] - Total protein Control (ab184247) staining at 1/1000 dilution.

All lanes:

Western blot - Anti-DRP1 (phospho S616) antibody [EPR27387-57] (<a href='/en-us/products/primary-antibodies/drp1-phospho-s616-antibody-epr27387-57-ab314755'>ab314755</a>) at 1/1000 dilution

Lane 1:

PC-12 (rat adrenal gland pheochromocytoma cell) whole cell lysate (untreated membrane) at 20 µg

Lane 2:

PC-12 whole cell lysate (phosphatase treated membrane) 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: 83 kDa

false

Exposure time: 180s

• WB

Supplier Data

Western blot - Anti-DRP1 antibody [EPR19274] (AB184247)

Blocking and diluting buffer and concentration : 5% NFDM/TBST. The identity of the lower MW band at approximately 20 kDa (in lane 2) is unknown. In western blot, anti-GAPDH antibody [EPR16891] - Loading Control (ab181602) staining at 1/200000 dilution. In western blot, anti-DRP1 antibody [EPR19274] - Total protein Control (ab184247) staining at 1/1000 dilution.

All lanes:

Western blot - Anti-DRP1 (phospho S616) antibody [EPR27387-57] (<a href='/en-us/products/primary-antibodies/drp1-phospho-s616-antibody-epr27387-57-ab314755'>ab314755</a>) at 1/1000 dilution

Lane 1:

Untreated NIH/3T3 (mouse embryonic fibroblast) whole cell lysate at 20 µg

Lane 2:

NIH/3T3 treated with /ml nocodazole for 17 hours 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: 83 kDa

false

Exposure time: 180s

• WB

Supplier Data

Western blot - Anti-DRP1 antibody [EPR19274] (AB184247)

Blocking and diluting buffer and concentration : 5% NFDM/TBST. Immunoprecipitation experiments were performed with an anti-6X His tag® antibody (ab213204) at 1 : 5000 dilution. In western blot, anti-DRP1 antibody [EPR19274] - Total protein Control (ab184247) staining at 1/1000 dilution.

All lanes:

Western blot - Anti-DRP1 (phospho S616) antibody [EPR27387-57] (<a href='/en-us/products/primary-antibodies/drp1-phospho-s616-antibody-epr27387-57-ab314755'>ab314755</a>) at 1/1000 dilution

Lane 1:

Untreated 293T cells transfected with a human DRP1 expression vector containi a myc-His-tag® lysate at 20 µg

Lane 2:

293T cells transfected with a human DRP1 expression vector containing a myc-His-tag®. Overexpressi cells were treated with 100/ml nocodazole for 17 hours whole cell lysate at 20 µg

Lane 3:

Untreated 293T cells transfected with a human DRP1(S616A mutant ) expression vector containi a myc-His-tag® lysate at 20 µg

Lane 4:

293T cells transfected with a human DRP1(S616A mutant) expression vector containing a myc-His-tag®. Overexpressi cells were treated with 100/ml nocodazole for 17 hours whole cell lysate at 20 µg

Secondary

All lanes:

Western blot - 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: 83 kDa

false

Exposure time: 3s