Anti-CCK4 antibody [EPR30150-524]

6 product images

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  Immunoprecipitation - Anti-CCK4 antibody [EPR30150-524] (ab323570)

  CCK4 was immunoprecipitated from 0.35 mg A431 (human epidermoid carcinoma epithelial cell) whole cell lysate with ab323570 at 1/30 dilution (2ug in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab323570 at 1/1000 dilution.

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

  All lanes: Immunoprecipitation - Anti-CCK4 antibody [EPR30150-524] (ab323570) at 1/1000 dilution

  Lane 1: A431 (human epidermoid carcinoma epithelial cell) whole cell lysate at 10 µg

  Lane 2: ab323570 at 1/30 IP in A431 (human epidermoid carcinoma epithelial cell) whole cell lysate at 10 µg

  Lane 3: Rabbit monoclonal IgG (Rabbit IgG, monoclonal [EPR25A] - Isotype Control ab172730) instead of ab323570 in A431 whole cell lysate at 10 µg

  Secondary

  All lanes: Immunoprecipitation - VeriBlot for IP Detection Reagent (HRP) (VeriBlot for IP Detection Reagent (HRP) ab131366) at 1/5000 dilution

  Observed band size: 110 kDa

  Exposure time: 180s

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  Immunocytochemistry/ Immunofluorescence - Anti-CCK4 antibody [EPR30150-524] (ab323570)

  Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized A431 (human epidermoid carcinoma epithelial cell) cells labelling CCK4 with ab323570 at 1/50 (10.12 ug/ml) dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/ 1000 2ug/ml dilution (Green).

  Confocal image showing mainly membranous staining in A431 cell line (shown in green). The counterstain was observed in magenta. Nuclear DNA was labelled with DAPI (shown in blue).
  
  Low expression: A549.
  
  Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

  Alexa Fluor® 594 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor® 594) was used to counterstain tubulin at 1/200 2.5ug/ml dilution (Magenta). The Nuclear counterstain was DAPI (Blue).

  Secondary antibody only control: Secondary antibody is Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/ 1000 2ug/ml dilution.

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  Western blot - Anti-CCK4 antibody [EPR30150-524] (ab323570)

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

  CCK4 is a glycoprotein of approximately 110 and 160 kDa and detected as 110kDa and 80 kDa band after treated with Deglycosylation Peptide:N-glycosidase F (PNGase F).

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

  All lanes: Western blot - Anti-CCK4 antibody [EPR30150-524] (ab323570) at 1/1000 dilution

  Lane 1: Untreated A431 (human epidermoid carcinoma epithelial cell) whole cell lysate at 20 µg

  Lane 2: A431 whole cell lysate treated with Protein Deglycosylation Peptide:N-glycosidase F (PNGase F) at 20 µg

  Secondary

  All lanes: Western blot - Goat Anti-Rabbit IgG H&L (HRP) (Goat Anti-Rabbit IgG H&L (HRP) ab97051) at 1/20000 dilution

  Observed band size: 110,160 kDa, 110, 80 kDa, 36 kDa

  Exposure time: 70s

• 

  Western blot - Anti-CCK4 antibody [EPR30150-524] (ab323570)

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

  Low expression: colon

  The identity of the bands lower than 75kDa are unknown.

  This blot of lane 1 was developed using a high-sensitivity ECL substrate, allowing for the detection of proteins in the mid-femtogram range.

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

  All lanes: Western blot - Anti-CCK4 antibody [EPR30150-524] (ab323570) at 1/1000 dilution

  Lane 1: Human lung tissue lysate at 20 µg

  Lane 2: Human colon tissue lysate at 20 µg

  Lane 3: Human breast cancer tissue lysate at 20 µg

  Secondary

  All lanes: Goat Anti-Rabbit IgG (HRP) with minimal cross-reactivity with human IgG at 1/2000 dilution

  Observed band size: 110 kDa, 36 kDa

  Exposure time: 180s

• 

  Western blot - Anti-CCK4 antibody [EPR30150-524] (ab323570)

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

  Low expression: A549, HT-1080(PMID: 25006253).

  The expression profile observed is consistent with what has been described in the literature(PMID: 25006253)

  This antibody can recognize both the full-length PTK7 and the N-terminal PTK7. The 160 kDa form may represent the full-length protein with glycosylation modifications, while the 110 kDa form likely corresponds to the N-terminal PTK7 glycosylation modification.

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

  All lanes: Western blot - Anti-CCK4 antibody [EPR30150-524] (ab323570) at 1/1000 dilution

  Lane 1: HCT 116 (human colorectal carcinoma epithelial cell) whole cell lysate at 20 µg

  Lane 2: HT-29 (human colorectal adenocarcinoma epithelial cell) whole cell lysate at 20 µg

  Lane 3: HeLa (human cervical adenocarcinoma epithelial cell) whole cell lysate at 20 µg

  Lane 4: A431 (human epidermoid carcinoma epithelial cell) whole cell lysate at 20 µg

  Lane 5: SW480 (human colorectal adenocarcinoma epithelial cell) whole cell lysate at 20 µg

  Lane 6: A549 (human lung carcinoma epithelial cell) whole cell lysate at 20 µg

  Lane 7: HT-1080 (human fibrosarcoma epithelial cell) whole cell lysate at 20 µg

  Secondary

  All lanes: Western blot - Goat Anti-Rabbit IgG H&L (HRP) (Goat Anti-Rabbit IgG H&L (HRP) ab97051) at 1/20000 dilution

  Observed band size: 110,160 kDa, 36 kDa

  Exposure time: 48s

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  Flow Cytometry - Anti-CCK4 antibody [EPR30150-524] (ab323570)

  Flow cytometric analysis of A431 (human epidermoid carcinoma epithelial cell) (Right) / A549 (human lung carcinoma epithelial cell) (Left) cells labelling CCK4 with ab323570 at 1/500 dilution (0.1ug) (Red) compared with a Rabbit monoclonal IgG (Rabbit IgG, monoclonal [EPR25A] - Isotype Control ab172730) (Black) isotype control and an unlabelled control (cells without incubation with primary antibody and secondary antibody).

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

  Gated on viable cells.
  
  Low expression: A549.