Anti-Clathrin heavy chain antibody [EPR24236-222]

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-Clathrin heavy chain antibody [EPR24236-222] (AB305259)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized HeLa (human cervical adenocarcinoma epithelial cell) cells labelling Clathrin heavy chain with ab305259 at 1/50 dilution, followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 dilution (Green). Confocal image showing punctuated cytoplasmic staining in HeLa cell line. The image was taken with a confocal microscope (Leica-Microsystems, TCS SP8). ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor® 594) was used to counterstain tubulin at 1/200 dilution (Red). The Nuclear counterstain was DAPI (Blue). Secondary antibody only control : Secondary antibody is ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 dilution.

• Flow Cyt (Intra)

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Flow Cytometry (Intracellular) - Anti-Clathrin heavy chain antibody [EPR24236-222] (AB305259)

Flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized HeLa (human cervix adenocarcinoma epithelial cell) cells labelling Clathrin heavy chain with ab305259 at 1/500 dilution (0.1ug) (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). A Goat Anti-Rabbit IgG (Alexa Fluor® 488, ab150081) at 1/2000 dilution was used as the secondary antibody.

• IP

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Immunoprecipitation - Anti-Clathrin heavy chain antibody [EPR24236-222] (AB305259)

Clathrin heavy chain was immunoprecipitated from 0.35 mg HeLa (human cervix adenocarcinoma epithelial cell) whole cell lysate 10 μg with ab305259 at 1/30 dilution (2μg in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab305259. VeriBlot for IP secondary antibody(HRP)(ab131366) was used at 1/5000 dilution. Lane 1 : HeLa (human cervix adenocarcinoma epithelial cell) whole cell lysate 10 μg Lane 2 : ab305259 IP in HeLa whole cell lysate Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab305259 in HeLa whole cell lysate Blocking and dilution buffer and concentration : 5% NFDM/TBST. Exposure time : 50 seconds

All lanes:

Immunoprecipitation - Anti-Clathrin heavy chain antibody [EPR24236-222] (ab305259) at 1/30 dilution

All lanes:

HeLa (human cervix adenocarcinoma epithelial cell)

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: 180 kDa

false

Exposure time: 50s

• WB

Supplier Data

Western blot - Anti-Clathrin heavy chain antibody [EPR24236-222] (AB305259)

Blocking and diluting buffer and concentration : 5% NFDM/TBST Exposure time : 37 seconds

All lanes:

Western blot - Anti-Clathrin heavy chain antibody [EPR24236-222] (ab305259) at 1/1000 dilution

Lane 1:

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

Lane 2:

293T (human embryonic kidney epithelial cell) 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 1/2000 dilution

Observed band size: 180 kDa

false

Exposure time: 37s