Anti-MUPP1 antibody [EPR26317-59]

13 product images

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  Immunoprecipitation - Anti-MUPP1 antibody [EPR26317-59] (ab302621)

  MUPP1 was immunoprecipitated from 0.35 mg C6 (rat glial tumor glial cell), whole cell lysate 10 µg with ab302621 at 1/30 dilution (2µg in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab302621 at 1/1000 dilution. VeriBlot for IP secondary antibody(HRP)(VeriBlot for IP Detection Reagent (HRP) ab131366) was used at 1/5000 dilution.
  

  Lane 1: C6 (rat glial tumor glial cell), whole cell lysate 10 µg

  Lane 2: ab302621 IP in C6 whole cell lysate

  Lane 3: Rabbit monoclonal IgG (Rabbit IgG, monoclonal [EPR25A] - Isotype Control ab172730) instead of ab302621 in C6 whole cell lysate

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

  This blot was developed using a high sensitivity ECL substrate. The high-sensitivity ECL substrate used allows for the detection of proteins in the mid-femtogram range.

  The bands beneath the target band are likely to be degraded target fragments.

  All lanes: Immunoprecipitation - Anti-MUPP1 antibody [EPR26317-59] (ab302621) at 1/30 dilution

  All lanes: C6 (rat glial tumor glial cell), whole cell lysate

  Secondary

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

  Developed using the ECL technique.

  Predicted band size: 219 kDa

  Exposure time: 58s

• 

  Western blot - Anti-MUPP1 antibody [EPR26317-59] (ab302621)

  Western blot: Anti-MPDZ antibody [EPR26317-59] (ab302621) staining at 1/1000 dilution, shown in green; Mouse anti-Alpha Tubulin [DM1A] (Anti-alpha Tubulin antibody [DM1A] - Loading Control ab7291) loading control staining at 1/20000 dilution, shown in magenta. In Western blot, ab302621 was shown to bind specifically to MPDZ. A band was observed at 268 kDa in wild-type A549 cell lysates with no signal observed at this size in MPDZ knockout cell line. To generate this image, wild-type and MPDZ knockout A549 cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3 % milk in TBS-0.1 % Tween® 20 (TBS-T) before incubation with primary antibodies overnight at 4 °C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Rabbit IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution.

  All lanes: Western blot - Anti-MUPP1 antibody [EPR26317-59] (ab302621) at 1/1000 dilution

  Lane 1: Wild-type A549 cell lysate at 20 µg

  Lane 2: MPDZ knockout A549 cell lysate at 20 µg

  Lane 3: SH-SY5Y cell lysate at 20 µg

  Lane 4: SW480 cell lysate at 20 µg

  Secondary

  All lanes: Goat anti-Rabbit IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution

  Performed under reducing conditions.

  Observed band size: 268 kDa

• 

  Flow Cytometry (Intracellular) - Anti-MUPP1 antibody [EPR26317-59] (ab302621)

  Flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized mIMCD3 (mouse inner medullary collecting duct epithelial cell) cells labelling MUPP1 with ab302621 at 1/500 dilution (0.1ug) (Red) (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) (Blue). A Goat Anti-Rabbit IgG (Alexa Fluor® 488, Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed ab150081) at 1/2000 dilution was used as the secondary antibody.

• 

  Immunohistochemistry (Frozen sections) - Anti-MUPP1 antibody [EPR26317-59] (ab302621)

  Immunohistochemical analysis of 4% PFA-fixed, 0.2% Triton X-100 permeabilized frozen Rat choroid plexus tissue labeling MUPP1 with ab302621 at 1/50 (10.92 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 at 1/1000 2 ug/mL dilution (Green). Positive staining on rat choroid plexus is observed. The nuclear counterstain was DAPI (Blue).

  Secondary antibody control: Secondary antibody is Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbedat 1/1000 2 ug/mL dilution.<\p>

  Heat mediated antigen retrieval using sodium citrate buffer (10mM citrate pH 6.0 + 0.05% Tween-20).

• 

  Immunohistochemistry (Frozen sections) - Anti-MUPP1 antibody [EPR26317-59] (ab302621)

  Immunohistochemical analysis of 4% PFA-fixed, 0.2% Triton X-100 permeabilized frozen Mouse choroid plexus tissue labeling MUPP1 with ab302621 at 1/50 (10.92 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 at 1/1000 2 ug/mL dilution (Green). Positive staining on mouse choroid plexus is observed. The nuclear counterstain was DAPI (Blue).

  Secondary antibody control: Secondary antibody is Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbedat 1/1000 2 ug/mL dilution.<\p>

  Heat mediated antigen retrieval using sodium citrate buffer (10mM citrate pH 6.0 + 0.05% Tween-20).

• 

  Flow Cytometry (Intracellular) - Anti-MUPP1 antibody [EPR26317-59] (ab302621)

  Flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized A549 (human lung carcinoma epithelial cell) cells labelling MUPP1 with ab302621 at 1/500 dilution (0.1ug) (Red) (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) (Blue). A Goat Anti-Rabbit IgG (Alexa Fluor® 488, Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed ab150081) at 1/2000 dilution was used as the secondary antibody.

• 

  Immunocytochemistry/ Immunofluorescence - Anti-MUPP1 antibody [EPR26317-59] (ab302621)

  Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized mIMCD3 (mouse inner medullary collecting duct epithelial cell) cells labelling MUPP1 with ab302621 at 1/50 (10.92 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 2 ug/ml dilution (Green). Confocal image showing membranous and cytoplasmic staining in mIMCD3 cell line is observed. 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 (Red). 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 2 ug/ml dilution.

• 

  Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-MUPP1 antibody [EPR26317-59] (ab302621)

  Immunohistochemical analysis of paraffin-embedded Rat skeletal muscle tissue labeling MUPP1 with ab302621 at 1/800 (0.683 ug/ml) followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection) was used. Negative control: no staining in rat skeletal muscle (PMID: 12706259). The section was incubated with ab302621 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) was used.
  Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins

• 

  Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-MUPP1 antibody [EPR26317-59] (ab302621)

  Immunohistochemical analysis of paraffin-embedded Mouse skeletal muscl tissue labeling MUPP1 with ab302621 at 1/800 (0.683 ug/ml) followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection) was used. Negative control: no staining in mouse skeletal muscle (PMID: 12706259). The section was incubated with ab302621 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) was used.
  Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins

• 

  Western blot - Anti-MUPP1 antibody [EPR26317-59] (ab302621)

  Blocking and diluting buffer and concentration: 5% NFDM/TBST
  The expression profile/ molecular weight observed is consistent with what has been described in the literature (PMID: 12403818).
  
  Low expression: skeletal muscle (PMID: 12706259)
  
  Lysate in lane 5 was freshly made and used for Western blotting immediately to minimize protein degradation.
  
  The bands beneath the target band are likely to be degraded target fragments.
  
  Samples are non-boiled as boiling may cause protein aggregates.
  Exposure time: Lane 1-4: 26 seconds
  Lane 5: 3 minutes

  All lanes: Western blot - Anti-MUPP1 antibody [EPR26317-59] (ab302621) at 1/1000 dilution

  Lane 1: Mouse brain tissue lysate 40 µg

  Lane 2: Mouse skeletal muscle tissue lysate 40 µg

  Lane 3: Rat brain tissue lysate 40 µg

  Lane 4: Rat skeletal muscle tissue lysate 40 µg

  Lane 5: Mouse testis tissue lysate 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: 280 kDa, 278 kDa

  Exposure time: 26s

• 

  Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-MUPP1 antibody [EPR26317-59] (ab302621)

  Immunohistochemical analysis of paraffin-embedded Rat choroid plexus tissue labeling MUPP1 with ab302621 at 1/800 (0.683 ug/ml) followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection) was used. Positive staining in rat choroid plexus (PMID: 30518636, PMID: 12706259 ). The section was incubated with ab302621 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) was used.
  Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins

• 

  Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-MUPP1 antibody [EPR26317-59] (ab302621)

  Immunohistochemical analysis of paraffin-embedded Mouse choroid plexus tissue labeling MUPP1 with ab302621 at 1/800 (0.683 ug/ml) followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection) was used. Positive staining in mouse choroid plexus (PMID: 30518636, PMID: 12706259). The section was incubated with ab302621 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) was used.
  Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins

• 

  Western blot - Anti-MUPP1 antibody [EPR26317-59] (ab302621)

  Blocking and diluting buffer and concentration: 5% NFDM/TBST
  The expression profile/ molecular weight observed is consistent with what has been described in the literature (PMID: 12403818).
  
  Lysates in lane1-4 were freshly made and used for Western blotting immediately to minimize protein degradation.
  
  The bands beneath the target band are likely to be degraded target fragments.
  
  Samples are non-boiled as boiling may cause protein aggregates.
  Exposure time: Lane 1-7: 3 minutes
  Lane 8: 26 seconds

  All lanes: Western blot - Anti-MUPP1 antibody [EPR26317-59] (ab302621) at 1/1000 dilution

  Lane 1: C6 (rat glial tumor glial cell), whole cell fresh lysate 20 µg

  Lane 2: Neuro-2a (mouse neuroblastoma neuroblast), whole cell fresh lysate 20 µg

  Lane 3: U-87 MG (human glioblastoma-astrocytoma epithelial cell), whole cell fresh lysate 20 µg

  Lane 4: SH-SY5Y (human neuroblastoma epithelial cell), whole cell fresh lysate 20 µg

  Lane 5: C6 (rat glial tumor glial cell), whole cell lysate 40 µg

  Lane 6: PC-12 (rat adrenal gland pheochromocytoma), whole cell lysate 40 µg

  Lane 7: human cerebellum tissue lysate 40 µg

  Lane 8: mIMCD3 (mouse inner medlary collecting duct epithelial cell), whole cell lysate 40 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: 280 kDa

  Exposure time: 3min