Anti-CD9 antibody [EPR23105-125] - BSA and Azide free

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CD9 antibody [EPR23105-125] - BSA and Azide free (AB263024)

Immunohistochemical analysis of paraffin-embedded Human cervical carcinoma tissue labeling CD9 with ab263019 at 1/1000 dilution followed by a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Positive staining on human cervical carcinoma. The section was incubated with ab263019 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 Rabbit specific IHC polymer detection kit HRP/DAB (ab209101).

Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution 2) for 20mins.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab263019).

• Flow Cyt

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Flow Cytometry - Anti-CD9 antibody [EPR23105-125] - BSA and Azide free (AB263024)

Flow cytometric analysis of Raji (human burkitt's lymphoma b lymphocyte, Left) / HCT 116 (human colorectal carcinoma epithelial cell, Right) cells labelling CD9 with ab263019 at UNABLE TO ENCODE THE VARIABLE PLEASE DO MANUALLY compared with a Rabbit monoclonal IgG (ab172730) / Black isotype control and an unlabelled control (cells without incubation with primary antibody and secondary antibody) (Blue). Goat anti rabbit IgG (Alexa Fluor® 488, ab150077) at 1/2000 was used as the secondary antibody. This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab263019).

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CD9 antibody [EPR23105-125] - BSA and Azide free (AB263024)

Immunohistochemical analysis of paraffin-embedded Human spleen tissue labeling CD9 with ab263019 at 1/1000 dilution followed by a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Positive staining on platelets of human spleen. The section was incubated with ab263019 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 Rabbit specific IHC polymer detection kit HRP/DAB (ab209101).

Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution 2) for 20 mins.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab263019).

• IP

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Immunoprecipitation - Anti-CD9 antibody [EPR23105-125] - BSA and Azide free (AB263024)

CD9 was immunoprecipitated from 0.35 mg HCT 116 (human colorectal carcinoma epithelial cell) whole cell lysate with ab263019 at 1/30 dilution (2ug in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab263019 at 1/500 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366) was used at 1/1000 dilution.

Lane 1 : HCT 116 (human colorectal carcinoma epithelial cell) whole cell lysate 10ug.

Lane 2 : ab263019 IP in HCT 116 whole cell lysate.

Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab263019 in HCT 116 whole cell lysate.

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

Exposure time : 10 seconds.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab263019).

All lanes:

Immunoprecipitation - Anti-CD9 antibody [EPR23105-125] (<a href='/en-us/products/primary-antibodies/cd9-antibody-epr23105-125-ab263019'>ab263019</a>)

Predicted band size: 25 kDa

Observed band size: 22 kDa

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• WB

Lab

Western blot - Anti-CD9 antibody [EPR23105-125] - BSA and Azide free (AB263024)

This data was developed using the same antibody clone in a different buffer formulation (ab263019).

Lanes 1 - 4 : Merged signal (red and green). Green - ab263019 observed at 18 kDa. Red - loading control ab7291 (Mouse anti-Alpha Tubulin [DM1A]) observed at 55 kDa.

ab263019 was shown to react with CD9 in wild-type HeLa cells in Western blot with loss of signal observed in CD9 knockout cell line ab255375 (CD9 knockout cell lysate ab263754). Wild-type HeLa and CD9 knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in fluorescent western blot (TBS-based) blocking solution before incubation with ab263019 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 h at room temperature before imaging.

All lanes:

Western blot - Anti-CD9 antibody [EPR23105-125] (<a href='/en-us/products/primary-antibodies/cd9-antibody-epr23105-125-ab263019'>ab263019</a>) at 1/1000 dilution

Lane 1:

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

Lane 2:

CD9 knockout HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate at 20 µg

Lane 2:

Western blot - Human CD9 knockout HeLa cell line (<a href='/en-us/products/cell-lines/human-cd9-knockout-hela-cell-line-ab255375'>ab255375</a>)

Lane 3:

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

Lane 4:

MCF7 (Human breast adenocarcinoma cell line) whole cell lysate at 20 µg

Predicted band size: 25 kDa

Observed band size: 18 kDa

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