Anti-Cyclophilin B antibody (ab16045)

Anti-Cyclophilin B antibody images

• 

  Immunocytochemistry/ Immunofluorescence - Anti-Cyclophilin B antibody (ab16045)

  ab16045 stained HeLa cells. The cells were 100% methanol fixed for 5 minutes at -20°C and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1hour at room temperature to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab16045 at 5µg/ml) overnight at +4°C. The secondary antibody (pseudo-colored green) was Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed (ab150081) used at a 1/1000 dilution for 1hour at room temperature. Alexa Fluor® 594 WGA was used to label plasma membranes (pseudo-colored red) at a 1/200 dilution for 1hour at room temperature. DAPI was used to stain the cell nuclei (pseudo-colored blue) at a concentration of 1.43µM for 1hour at room temperature.

• 

  Western blot - Cyclophilin B antibody (ab16045)
  All lanes : Anti-Cyclophilin B antibody (ab16045) at 1 µg/ml
  
  Lane 1 : Rat Liver
  Lane 2 : Mouse 3T3
  Lane 3 : Dog
  Lane 4 : Chicken
  
  Lysates/proteins at 20 µg per lane.
  
  Secondary
  Goat Anti-Rabbit IgG H&L (HRP) (ab6721) at 1/5000 dilution
  
  Performed under reducing conditions.
  
  Predicted band size : 21 kDa
  Observed band size : 25 kDa (why is the actual band size different from the predicted?)
  
  
  Exposure time : 30 seconds
• 

  Western blot - Anti-Cyclophilin B antibody (ab16045)
  All lanes : Anti-Cyclophilin B antibody (ab16045) at 1 µg/ml
  
  Lane 1 : HeLa nuclear lysate
  Lane 2 : HeLa whole cell lysate
  Lane 3 : A431 whole cell lysate
  Lane 4 : Jurkat whole cell lysate
  Lane 5 : HEK293 whole cell lysate
  Lane 6 : HeLa nuclear lysate with Human Cyclophilin B peptide (ab16277) at 1 µg/ml
  Lane 7 : HeLa whole cell lysate with Human Cyclophilin B peptide (ab16277) at 1 µg/ml
  Lane 8 : A431 whole cell lysate with Human Cyclophilin B peptide (ab16277) at 1 µg/ml
  Lane 9 : Jurkat whole cell lysate with Human Cyclophilin B peptide (ab16277) at 1 µg
  Lane 10 : HEK293 whole cell lysate with Human Cyclophilin B peptide (ab16277) at 1 µg/ml
  
  Lysates/proteins at 20 µg per lane.
  
  Secondary
  Goat Anti-Rabbit IgG H&L (HRP) (ab6721) at 1/5000 dilution
  
  Performed under reducing conditions.
  
  Predicted band size : 21 kDa
  Observed band size : 21 kDa
  
  
  Exposure time : 30 seconds
• 

  Immunoprecipitation - Anti-Cyclophilin B antibody (ab16045)
  Cyclophilin B was immunoprecipitated using 0.5mg Hela whole cell extract, 5µg of Rabbit polyclonal to Cyclophilin B and 50µl of protein G magnetic beads (+). No antibody was added to the control (-).
  The antibody was incubated under agitation with Protein G beads for 10min, Hela whole cell extract lysate diluted in RIPA buffer was added to each sample and incubated for a further 10min under agitation.
  Proteins were eluted by addition of 40µl SDS loading buffer and incubated for 10min at 70^oC; 10µl of each sample was separated on a SDS PAGE gel, transferred to a nitrocellulose membrane, blocked with 5% BSA and probed with ab16045.
  Secondary: Clean blot (HRP conjugate) at 1/1000 dilution.
  Band: 21kDa: Cyclophilin B.
• 

  Immunocytochemistry/ Immunofluorescence - Cyclophilin B antibody (ab16045)

  ICC/IF image of ab16045 stained NIH/3T3 cells. The cells were methanol fixed (5 min) and incubated with the antibody (ab16045, 1µg/ml) for 1h at room temperature. The secondary antibody (green) was Alexa Fluor® 488 goat anti-rabbit IgG (H+L) used at a 1/1000 dilution for 1h. Image-iT^TM FX Signal Enhancer was used as the primary blocking agent, 5% BSA (in TBS-T) was used for all other blocking steps. DAPI was used to stain the cell nuclei (blue). Alexa Fluor® 594 WGA was used to label plasma membranes (red).

• 

  Immunocytochemistry/ Immunofluorescence - Cyclophilin B antibody (ab16045)This image is courtesy of an Abreview submitted by Dr Kirk McManus
  ab16045 (1/1000) staining Cyclophilin B in assynchronous HeLa cells (green). Cells were fixed with Paraformaldehyde and counter-stained with DAPI in order to highlight the nucleus (red). Please refer to abreview for further experimental details.

  See Abreview

• 

  Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Cyclophilin B antibody (ab16045)

  Image courtesy of Human Protein Atlas

  ab16045 staining in human placenta, showing staining of the cytrotrophoblasts. Paraffin embedded placental tissue was incubated with ab16045 (1:14,000 dilution) for 30 mins at room temperature. Antigen retrieval was performed by heat induction in citrate buffer pH 6. ab16045 was tested in a tissue microarray (TMA) containing a wide range of normal and cancer tissues as well as a cell microarray consisting of a range of commonly used, well characterised human cell lines. Further results for this antibody can be found at www.proteinatlas.org

  .

• 

  Western blot - Cyclophilin B antibody (ab16045)Image courtesy of Dr M Schrader by Abreview.
  All lanes : Anti-Cyclophilin B antibody (ab16045) at 1/1000 dilution
  
  Lane 1 : Whole cell lysate prepared from rat pancreatic AR42J cells, which were treated with 10nM dexamethasone for 48 hours.
  Lane 2 : Whole cell lysate for negative control, prepared from rat pancreatic AR42J cells (specific knock down of cyclophilin B/PpiB by siRNA), which were treated with 10nM dexamethasone for 48 hours.
  
  Secondary
  Goat-anti-Rabbit HRP-conjugated polyclonal at 1/2000 dilution
  Developed using the ECL technique
  
  Predicted band size : 21 kDa
  Observed band size : 23 kDa (why is the actual band size different from the predicted?)
  
  
  Exposure time : 1 minute

  Image courtesy of Dr M Schrader by Abreview.

  Primary antibody incubated for 12 hours at 4°C.
  Blocking step performed using 5% milk, 1 hour at 20°C.

  See Abreview

• 

  Western blot - Anti-Cyclophilin B antibody (ab16045)
  Anti-Cyclophilin B antibody (ab16045) at 0.5 µg/ml + Recombinant Human Cyclophilin B protein (ab88801) at 0.01 µg
  
  Secondary
  Goat Anti-Rabbit IgG H&L (HRP) preadsorbed (ab97080) at 1/5000 dilution
  Developed using the ECL technique
  
  Performed under reducing conditions.
  
  Predicted band size : 21 kDa
  
  
  Exposure time : 30 seconds
• 

  Western blot - Anti-Cyclophilin B antibody (ab16045)
  
  Developed using the ECL technique
  
  Performed under reducing conditions.
  
  Predicted band size : 21 kDa