All lanes : Anti-Integrin alpha V antibody [EPR16800] (ab179475) at 1/50000 dilution

Lane 1 : HUVEC (Human umbilical vein endothelial cell line) whole cell lysates
Lane 2 : HT-29 (Human colorectal adenocarcinoma cells) whole cell lysates
Lane 3 : A549 (Human lung carcinoma) whole cell lysates

Lysates/proteins at 20 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG, (H+L),Peroxidase conjugated at 1/1000 dilution

Predicted band size: 116 kDa
Observed band size: 125 kDa why is the actual band size different from the predicted?

Blocking/dilution buffer: 5% NFDM/TBST.

ab179475 can recognize 135kDa full length Integrin alpha V and 125kDa heavy chain. The 125 kDa band is Integrin alpha V heavy chain.

Immunohistochemical analysis of paraffin-embedded human transitional cell carcinoma of bladder tissue labeling Integrin alpha V with ab179475 at 1/500 dilution followed by Goat Anti-Rabbit HRP (IgG H&L) (ab97051) secondary antibody at 1/500 dilution.

Membrane and weak cytoplasm staining on human transitional cell carcinoma of bladder is observed. Counterstained with hematoxylin.

Negative control: Using 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.

Integrin alpha V was immunoprecipitated from 1 mg of A549 (Human lung carcinoma) whole cell extract with ab179475 at 1/40 dilution. Western blot analysis was performed from the immunoprecipitate using ab179475 at 1/1000 dilution. Anti-Rabbit IgG (HRP), specific to the non-reduced form of IgG, was used as secondary antibody at 1/1500 dilution.

Lane 1: A549 whole cell extract.

Lane 2: PBS instead of A549 whole cell extract.

Blocking/Dilution buffer: 5% NFDM/TBST.

ab179475 can recognize 135kDa full length Integrin alpha V and 125kDa heavy chain. The 125 kDa band is Integrin alpha V heavy chain.

Immunofluorescent analysis of 100% methanol-fixed, 0.1% Triton X-100 permeabilized A549 (Human lung carcinoma) cells labeling Integrin alpha V with ab179475 at 1/500 dilution, followed by Goat anti-rabbit Alexa Fluor^® 488 (IgG) (ab150077) secondary antibody at 1/400 dilution (green).

Confocal image showing membrane and cytoplasm staining on A549 cell line. The nuclear counterstain is DAPI (blue). Tubulin is detected with ab7291 (anti-Tubulin mouse mAb) at 1/500 dilution and ab150120 (goat anti-mouse AlexaFluor^®594 secondary antibody) at 1/500 dilution (red).

The negative controls are as follows:

-ve control 1: ab179475 at 1/500 dilution followed by ab150120 (AlexaFluor^®594 Goat anti-Mouse secondary) at 1/500 dilution.

-ve control 2: ab7291 (anti-Tubulin mouse mAb) at 1/500 dilution followed by ab150077 (Alexa Fluor^®488 Goat Anti-Rabbit IgG H&L) at 1/400 dilution.

All lanes : Anti-Integrin alpha V antibody [EPR16800] (ab179475) at 1/5000 dilution

Lane 1 : Human fetal kidney lysates
Lane 2 : Human fetal brain lysates

Lysates/proteins at 10 µg per lane.

Secondary
All lanes : Anti-Rabbit IgG (HRP), specific to the non-reduced form of IgG at 1/1000 dilution

Predicted band size: 116 kDa
Observed band size: 125 kDa why is the actual band size different from the predicted?

Blocking/dilution buffer: 5% NFDM/TBST.

ab179475 can recognize 135kDa full length Integrin alpha V and 125kDa heavy chain. The 125 kDa band is Integrin alpha V heavy chain.

Immunohistochemical analysis of paraffin-embedded human kidney tissue labeling Integrin alpha V with ab179475 at 1/500 dilution followed by Goat Anti-Rabbit HRP (IgG H&L) (ab97051) secondary antibody at 1/500 dilution.

Membrane and cytoplasm staining on human kidney tubules is observed. Counterstained with hematoxylin.

Negative control: Using 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.

All lanes : Anti-Integrin alpha V antibody [EPR16800] (ab179475) at 1/5000 dilution

Lane 1 : Mouse brain lysates
Lane 2 : Mouse kidney lysates
Lane 3 : Mouse spleen lysates
Lane 4 : Rat brain lysates
Lane 5 : Rat kidney lysates
Lane 6 : C6 (Rat glial tumor cells) whole cell lysates
Lane 7 : NIH/3T3 (Mouse embyro fibroblast cells) whole cell lysates
Lane 8 : Raji (Human Burkitt's lymphoma cell line) whole cell lysates

Lysates/proteins at 10 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG, (H+L),Peroxidase conjugated at 1/1000 dilution

Predicted band size: 116 kDa
Observed band size: 125,135 kDa why is the actual band size different from the predicted?

Blocking/dilution buffer: 5% NFDM/TBST.

ab179475 can recognize 135kDa full length Integrin alpha V and 125kDa heavy chain. The 125 kDa band is Integrin alpha V heavy chain.

Immunohistochemical analysis of paraffin-embedded mouse kidney tissue labeling Integrin alpha V with ab179475 at 1/500 dilution followed by Goat Anti-Rabbit HRP (IgG H&L) (ab97051) secondary at 1/500 dilution.

Membrane and cytoplasm staining on mouse kidney tubule and glomerulus is observed. Counterstained with hematoxylin.

Negative control: Using 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.

All lanes : Anti-Integrin alpha V antibody [EPR16800] (ab179475) at 1/2000 dilution (8 hours at 4°C)

All lanes : Mouse skin whole cell lysate

Lysates/proteins at 40 µg per lane.

Secondary
All lanes : Goat anti-Rabbit IgG(H+L)-HRP at 1/5000 dilution

Developed using the ECL technique.

Predicted band size: 116 kDa


Exposure time: 25 seconds

Diluent: 2% BSA in 1X TBST.

Blocked using 5% BSA for 1 hour at RT.

 

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