Anti-CD9 antibody [EPR27551-92]

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CD9 antibody [EPR27551-92] (AB307085)

Immunohistochemical analysis of paraffin-embedded Rat kidney tissue labeling CD9 with ab307085 at 1/2000 (0.228 µg/ml) followed by a LeicaDS9800 (Bond™ Polymer Refine Detection) was used at Ready to use dilution. Membranous staining in rat kidney. The section was incubated with ab307085 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 LeicaDS9800 (Bond™ Polymer Refine Detection) was used at Ready to use dilution. Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CD9 antibody [EPR27551-92] (AB307085)

Immunohistochemical analysis of paraffin-embedded Mouse kidney tissue labeling CD9 with ab307085 at 1/2000 (0.228 µg/ml) followed by a LeicaDS9800 (Bond™ Polymer Refine Detection) was used at Ready to use dilution. Positive staining in mouse kidney. The section was incubated with ab307085 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 LeicaDS9800 (Bond™ Polymer Refine Detection) was used at Ready to use dilution. Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CD9 antibody [EPR27551-92] (AB307085)

Immunohistochemical analysis of paraffin-embedded Mouse cerebrum tissue labeling CD9 with ab307085 at 1/2000 (0.228 µg/ml) followed by a LeicaDS9800 (Bond™ Polymer Refine Detection) was used at Ready to use dilution. Positive staining in mouse cerebrum. The section was incubated with ab307085 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 LeicaDS9800 (Bond™ Polymer Refine Detection) was used at Ready to use dilution. Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CD9 antibody [EPR27551-92] (AB307085)

Immunohistochemical analysis of paraffin-embedded Mouse lung cancer tissue labeling CD9 with ab307085 at 1/2000 (0.228 µg/ml) followed by a LeicaDS9800 (Bond™ Polymer Refine Detection) was used at Ready to use dilution. Membranous staining in mouse lung cancer. The section was incubated with ab307085 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 LeicaDS9800 (Bond™ Polymer Refine Detection) was used at Ready to use dilution. Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CD9 antibody [EPR27551-92] (AB307085)

Immunohistochemical analysis of paraffin-embedded Rat cerebrum tissue labeling CD9 with ab307085 at 1/2000 (0.228 µg/ml) followed by a LeicaDS9800 (Bond™ Polymer Refine Detection) was used at Ready to use dilution. Positive staining in rat cerebrum. The section was incubated with ab307085 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 LeicaDS9800 (Bond™ Polymer Refine Detection) was used at Ready to use dilution. Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins

• IP

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Immunoprecipitation - Anti-CD9 antibody [EPR27551-92] (AB307085)

CD9 was immunoprecipitated from 0.35 mg RAW 264.7 (mouse Abelson murine leukemia virus-induced tumor macrophage) whole cell lysate 10 μg with ab307085 at 1/30 dilution (2μg in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab307085 at 1/1000 dilution. VeriBlot for IP secondary antibody(HRP)(ab131366) was used at 1/5000 dilution. Lane 1 : RAW 264.7 (mouse Abelson murine leukemia virus-induced tumor macrophage) whole cell lysate 10 μg Lane 2 : ab307085 IP in RAW 264.7 whole cell lysate Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab307085 in RAW 264.7 whole cell lysate Blocking and dilution buffer and concentration : 5% NFDM/TBST. Exposure time : 10 seconds

All lanes:

Immunoprecipitation - Anti-CD9 antibody [EPR27551-92] (ab307085) at 1/30 dilution

All lanes:

RAW 264.7 (mouse Abelson murine leukemia virus-induced tumor macrophage)

Secondary

All lanes:

Immunoprecipitation - VeriBlot for IP Detection Reagent (HRP) (<a href='/en-us/products/reagents/veriblot-for-ip-detection-reagent-hrp-ab131366'>ab131366</a>) at 1/5000 dilution

Observed band size: 23 kDa

false

Exposure time: 10s

• WB

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Western blot - Anti-CD9 antibody [EPR27551-92] (AB307085)

Blocking and diluting buffer and concentration : Intercept® (TBS) Blocking Buffer diluted with an equal volume of 0.1% TBS The samples were run on a Bis-Tris gel. Performed under reducing conditions. False colour image of Western blot : Anti-CD9 antibody [EPR27551-92] (ab307085) staining at 1/1000 dilution, shown in green; Mouse anti-GAPDH antibody [6C5] (ab8245) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab307085 was shown to bind specifically to CD9. A band was observed at 23 kDa in wild-type HeLa cell lysates whereas no signal observed at this size in CD9 knockout cell line ab255375 (knockout cell lysate ab263754). To generate this image, wild-type and CD9 knockout HeLa cell lysates were analyzed. First, samples were run on an SDS-PAGE gel then transferred onto an immobilon-FL PVDF membrane. Membranes were blocked in in Intercept® (TBS) Blocking Buffer diluted with an equal volume of 0.1% TBS 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 (IRDye® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (ab216776) at 1/20000 dilution.

All lanes:

Western blot - Anti-CD9 antibody [EPR27551-92] (ab307085) at 1/1000 dilution

Lane 1:

Wild-type HeLa (human cervical adenocarcinoma epithelial cell) whole cell lysate at 20 µg

Lane 2:

CD9 knockout HeLa whole cell lysate at 20 µg

Lane 3:

MCF7 (human breast adenocarcinoma epithelial cell) whole cell lysate at 20 µg

Lane 4:

RAW 264.7 (mouse Abelson murine leukemia virus-induced tumor macrophage) whole cell lysate at 20 µg

Secondary

Lanes 1 - 4:

Western blot - Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-irdye-800cw-preadsorbed-ab216773'>ab216773</a>) at 1/10000 dilution

Lanes 1 - 4:

Western blot - Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (<a href='/en-us/products/secondary-antibodies/goat-mouse-igg-h-l-irdye-680rd-preadsorbed-ab216776'>ab216776</a>) at 1/10000 dilution

Observed band size: 23 kDa

false

• WB

Supplier Data

Western blot - Anti-CD9 antibody [EPR27551-92] (AB307085)

Blocking and diluting buffer and concentration : 5% NFDM/TBST Negative controls : K-562, Raji (PMID : 17407154 ; PMID : 8921952). Exposure time : 70 seconds

All lanes:

Western blot - Anti-CD9 antibody [EPR27551-92] (ab307085) at 1/1000 dilution

Lane 1:

HCT 116 (human colorectal carcinoma epithelial cell) whole cell lysate at 20 µg

Lane 2:

K-562 (human chronic myelogenous leukemia lymphoblast) whole cell lysate at 20 µg

Lane 3:

Raji (human Burkitts lymphoma B lymphocyte) whole cell lysate at 20 µg

Lane 4:

Mouse brain tissue lysate at 20 µg

Lane 5:

Mouse kidney tissue lysate at 20 µg

Lane 6:

C6 (rat glial tumor glial cell) whole cell lysate at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/100000 dilution

Observed band size: 23 kDa

false

Exposure time: 70s