Anti-Caveolin-1 抗体 [E249] - Caveolae Marker

• Flow Cyt

Supplier Data

Flow Cytometry - Anti-Caveolin-1 antibody [E249] - Caveolae Marker (AB32577)

Flow cytometric analysis of 4% Paraformaldehyde fixed 90% Methanol permeabilized A431 (Human epidermoid carcinoma epithelial cell) cells labelling Caveolin-1 with ab32577 at 1/20 dilution (10 μg/ml) compared with a Rabbit monoclonal IgG (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, ab150077) at 1/2000 was used as the secondary antibody.

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Caveolin-1 antibody [E249] - Caveolae Marker (AB32577)

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of rat spleen tissue sections labelling Caveolin-1 with purified ab32577 at 1/2000 dilution (0.06 μg/ml). Heat mediated antigen retrieval was performed using Heat mediated antigen retrieval using Bond™ Epitope Retrieval Solution 2 (pH 9.0) . Tissue was counterstained with Hematoxylin. Rabbit specific IHC polymer detection kit HRP/DAB (ab209101) secondary antibody was used at 1 : 0 dilution. PBS instead of the primary antibody was used as the negative control. The immunostaining was performed on a Leica Biosystems BOND® RX instrument.

• WB

Lab

Western blot - Anti-Caveolin-1 antibody [E249] - Caveolae Marker (AB32577)

ab32577 was shown to react with CAV1 in wild-type U-87 MG cells in Western blot with loss of signal observed in CAV1 knockout cell line ab306811. Wild-type U-87 MG and CAV1 knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in 5% milk in TBST for 1 hr before incubation with ab32577 overnight at 4 °C at a 1/1000 dilution. Blots were incubated with secondary antibodies at 0.2 µg/mL before imaging.

This data was provided by YCharOS Inc., an open science company with the mission of characterizing commercially available antibody reagents for all human proteins. Abcam and YCharOS are working together to help address the reproducibility crisis by enabling the life science community to better evaluate commercially available antibodies.

All lanes:

Western blot - Anti-Caveolin-1 antibody [E249] - Caveolae Marker (ab32577)

Lane 1:

Wild-type U-87 MG lysate at 20 µg

Lane 2:

CAV1 knock-out U-87 MG lysate at 20 µg

Lane 2:

Western blot - Human CAV1 knockout U-87 MG cell line (ab306811)

Observed band size: 20-22 kDa

false

• WB

Lab

Western blot - Anti-Caveolin-1 antibody [E249] - Caveolae Marker (AB32577)

Lanes 1 - 4 : Merged signal (red and green). Green - ab32577 observed at 20 kDa. Red - loading control, ab8245 observed at 37 kDa.

ab32577 was shown to react with Caveolin-1 in wild-type HeLa cells. Loss of signal was observed when knockout cell line ab255371 (knockout cell lysate ab263806) was used. Wild-type and Caveolin-1 knockout samples were subjected to SDS-PAGE. ab32577 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated overnight at 4°C at 1 in 1000 dilution and 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preadsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.

All lanes:

Western blot - Anti-Caveolin-1 antibody [E249] - Caveolae Marker (ab32577) at 1/1000 dilution

Lane 1:

A431 cell lysate at 20 µg

Lane 2:

A549 cell lysate at 20 µg

Lane 2:

Western blot - Human CAV1 (Caveolin-1) knockout HeLa cell line (<a href='/products/cell-lines/human-cav1-caveolin-1-knockout-hela-cell-line-ab255371'>ab255371</a>)

Lane 3:

Wild-type HeLa cell lysate at 20 µg

Lane 4:

CAV1 knockout HeLa cell lysate at 20 µg

Secondary

All lanes:

Western blot - Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (<a href='/products/secondary-antibodies/goat-rabbit-igg-h-l-irdye-800cw-preadsorbed-ab216773'>ab216773</a>) at 1/20000 dilution

Predicted band size: 20 kDa

Observed band size: 20 kDa,37 kDa

false

• WB

Lab

Western blot - Anti-Caveolin-1 antibody [E249] - Caveolae Marker (AB32577)

Lanes 1 - 4 : Merged signal (red and green). Green - ab32577 observed at 20 kDa. Red - loading control, ab7291 (Mouse anti-Alpha Tubulin [DM1A] observed at 55kDa.

ab32577 was shown to react with Caveolin-1 in wild-type A431 cells in western blot. Loss of signal was observed when CAV1 knockout sample was used. A431 wild-type and CAV1 knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in 3% milk in TBS-T (0.1% Tween^®) before incubation with ab32577 and ab7291 (Mouse anti-Alpha Tubulin [DM1A] overnight at 4°C at a 1 in 1000 dilution and a 1 in 20000 dilution respectively. Blots were incubated with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preabsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.

All lanes:

Western blot - Anti-Caveolin-1 antibody [E249] - Caveolae Marker (ab32577) at 1/1000 dilution

Lane 1:

Wild-type A-431 (Human epidermoid carcinoma cell line) whole cell lysate at 20 µg

Lane 2:

CAV1 knockout A-431 (Human epidermoid carcinoma cell line) whole cell lysate at 20 µg

Lane 2:

Western blot - Human CAV1 (Caveolin-1) knockout A-431 cell line (<a href='/products/cell-lines/human-cav1-caveolin-1-knockout-a-431-cell-line-ab269583'>ab269583</a>)

Lane 3:

A549 (Human lung carcinoma cell line) whole cell lysate at 20 µg

Lane 4:

HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate at 20 µg

Predicted band size: 20 kDa

Observed band size: 20 kDa

false

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-Caveolin-1 antibody [E249] - Caveolae Marker (AB32577)

Immunocytochemistry analysis of Jurkat (Human lung carcinoma epithelial cell) cells labeling Caveolin-1 with purified ab32577 at 1/50 dilution (2.3 μg/mL). Cells were fixed in 4% Paraformaldehyde and permeabilized with 0.1% tritonX-100. Cells were counterstained with ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594) 1/200 (2.5 μg/mL). Goat anti rabbit IgG (Alexa Fluor^® 488, ab150078) was used as the secondary antibody at 1/1000 (2 μg/mL) dilution. DAPI (blue) was used as nuclear counterstain. PBS instead of the primary antibody was used as the secondary antibody only control.

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Caveolin-1 antibody [E249] - Caveolae Marker (AB32577)

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human tonsil tissue sections labelling Caveolin-1 with purified ab32577 at 1/2000 dilution (0.06 μg/ml). Heat mediated antigen retrieval was performed using Heat mediated antigen retrieval using Bond™ Epitope Retrieval Solution 2 (pH 9.0) . Tissue was counterstained with Hematoxylin. Rabbit specific IHC polymer detection kit HRP/DAB (ab209101) secondary antibody was used at 1 : 0 dilution. PBS instead of the primary antibody was used as the negative control. The immunostaining was performed on a Leica Biosystems BOND® RX instrument.

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Caveolin-1 antibody [E249] - Caveolae Marker (AB32577)

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of mouse lung tissue sections labelling Caveolin-1 with purified ab32577 at 1/2000 dilution (0.06 μg/ml). Heat mediated antigen retrieval was performed using Heat mediated antigen retrieval using Bond™ Epitope Retrieval Solution 2 (pH 9.0) . Tissue was counterstained with Hematoxylin. Rabbit specific IHC polymer detection kit HRP/DAB (ab209101) secondary antibody was used at 1 : 0 dilution. PBS instead of the primary antibody was used as the negative control. The immunostaining was performed on a Leica Biosystems BOND® RX instrument.

• ICC/IF

Unknown

Immunocytochemistry/ Immunofluorescence - Anti-Caveolin-1 antibody [E249] - Caveolae Marker (AB32577)

ab32577 staining Caveolin-1 in wild-type HeLa cells (top panel) and CAV1 knockout HeLa cells (ab255371) (bottom panel). The cells were fixed with 100% methanol (5 min) then permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated with ab32577 at 1/200 dilution and ab7291 (Mouse monoclonal to alpha Tubulin) at 1/1000 dilution overnight at 4°C followed by a further incubation at room temperature for 1h with a goat secondary antibody to rabbit IgG (Alexa Fluor^® 488) (ab150081) at 2 μg/ml (shown in green) and a goat secondary antibody to mouse IgG (Alexa Fluor^® 594) (ab150120) at 2 μg/ml (shown in red). Nuclear DNA was labelled in blue with DAPI.
Image was taken with a confocal microscope (Leica-Microsystems TCS SP8).

• WB

Lab

Western blot - Anti-Caveolin-1 antibody [E249] - Caveolae Marker (AB32577)

The two isoforms of Caveolin-1 have been described in the literatures (PMID : 12816877, 11748292 and 14992406). We are unsure about the nature of the 250kDa bands.

All lanes:

Western blot - Anti-Caveolin-1 antibody [E249] - Caveolae Marker (ab32577) at 1/1000 dilution

Lane 1:

Mouse heart lysate at 20 µg

Lane 2:

Mouse skeletal muscle lysate at 20 µg

Lane 3:

Rat heart lysate at 20 µg

Lane 4:

Rat skeletal muscle lysate at 20 µg

Lane 5:

C2C12 (Mouse myoblasts myoblast) whole cell lysate at 20 µg

Lane 6:

NIH/3T3 (Mouse embryonic fibroblast) whole cell lysate at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/20000 dilution

Predicted band size: 20 kDa

Observed band size: 17 kDa,23 kDa

false

• WB

Lab

Western blot - Anti-Caveolin-1 antibody [E249] - Caveolae Marker (AB32577)

The two isoforms of Caveolin-1 have been described in the literatures (PMID : 12816877, 11748292 and 14992406). We are unsure about the nature of the 250kDa bands.

All lanes:

Western blot - Anti-Caveolin-1 antibody [E249] - Caveolae Marker (ab32577) at 1/1000 dilution

Lane 1:

A431 (Human epidermoid carcinoma epithelial cell) whole cell lysate at 20 µg

Lane 2:

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

Lane 3:

HaCaT (Human skin keratinocyte) whole cell lysate at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/20000 dilution

Predicted band size: 20 kDa

Observed band size: 17 kDa,23 kDa

false