重组Anti-CD45抗体[EPR20033]

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CD45 antibody [EPR20033] (AB208022)

Immunohistochemical analysis of paraffin-embedded human skeletal muscle tissue labeling CD45 with ab208022 at 1/5000 (0.14 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Negative control : no staining in human skeletal muscle tissue.
The section was incubated with ab208022 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 LeicaDS9800 (Bond™ Polymer Refine Detection).

Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins.

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CD45 antibody [EPR20033] (AB208022)

Immunohistochemical analysis of paraffin-embedded human kidney tissue labeling CD45 with ab208022 at 1/5000 (0.14 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Negative control : no staining in human kidney tissue.
The section was incubated with ab208022 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 LeicaDS9800 (Bond™ Polymer Refine Detection).

Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins.

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CD45 antibody [EPR20033] (AB208022)

Immunohistochemical analysis of paraffin-embedded human tonsil tissue labeling CD45 with ab208022 at 1/5000 (0.14 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining in human tonsil tissue.
The section was incubated with ab208022 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 LeicaDS9800 (Bond™ Polymer Refine Detection).

Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins.

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CD45 antibody [EPR20033] (AB208022)

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of Mouse spleen tissue sections labeling CD45 with purified ab208022 at 1/5000 dilution (0.14 ?g/ml). Heat mediated antigen retrieval was performed. ImmunoHistoProbe one step HRP Polymer (ready to use) was used as the secondary antibody. Negative control : PBS instead of the primary antibody. Hematoxylin was used as a counterstain.

• IP

Supplier Data

Immunoprecipitation - Anti-CD45 antibody [EPR20033] (AB208022)

CD45 was immunoprecipitated from 0.35 mg of J774A.1 (Mouse reticulum cell sarcoma macrophage cell line) whole cell lysate with ab208022 at 1/30 dilution.

Western blot was performed from the immunoprecipitate using ab208022 at 1/1000 dilution.

VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/1000 dilution.

Lane 1 : J774A.1 whole cell lysate 10μg (Input).

Lane 2 : ab208022 IP in J774A.1 whole cell lysate.

Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab208022 in J774A.1 whole cell lysate.

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

Exposure time : 3 minutes.

All lanes:

Immunoprecipitation - Anti-CD45 antibody [EPR20033] (ab208022)

Predicted band size: 147 kDa

false

• WB

Lab

Western blot - Anti-CD45 antibody [EPR20033] (AB208022)

False colour image of Western blot : Anti-CD45 antibody [EPR20033] staining at 1/1000 dilution, shown in green; Mouse anti-Alpha Tubulin [DM1A] (ab7291) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab208022 was shown to bind specifically to CD45. A band was observed at 160-220 kDa in wild-type Jurkat cell lysates with no signal observed at this size in PTPRC CRISPR-Cas9 edited cell line ab274932 (CRISPR-Cas9 edited cell lysate ab274990). The band observed in the CRISPR-Cas9 edited lysate lane below 160-220 kDa is likely to represent a truncated form of CD45. This has not been investigated further and the functional properties of the gene product have not been determined. To generate this image, wild-type and PTPRC CRISPR-Cas9 edited Jurkat cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 5 % milk in TBS-0.1 % Tween^® 20 (TBS-T) 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-CD45 antibody [EPR20033] (ab208022) at 1/1000 dilution

Lane 1:

Wild-type Jurkat cell lysate at 20 µg

Lane 2:

PTPRC CRISPR-Cas9 edited Jurkat cell lysate at 20 µg

Lane 3:

THP-1 cell lysate at 20 µg

Lane 4:

MCF7 cell lysate at 20 µg

Predicted band size: 147 kDa

Observed band size: 160-220 kDa

false

• WB

Supplier Data

Western blot - Anti-CD45 antibody [EPR20033] (AB208022)

Blocking/Dilution buffer : 5% NFDM/TBST.

293T and MCF7 are reported to be CD45 negative cell lines (PMID : 22978632& 24602188).

All lanes:

Western blot - Anti-CD45 antibody [EPR20033] (ab208022) at 1/1000 dilution

Lane 1:

Jurkat (Human T cell leukemia cell line from peripheral blood) whole cell lysate at 20 µg

Lane 2:

293T (Human epithelial cell line from embryonic kidney) whole cell lysate at 20 µg

Lane 3:

MCF7 (Human breast adenocarcinoma cell line) 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 1/100000 dilution

Predicted band size: 147 kDa

Observed band size: 240 kDa

false

Exposure time: 3min

• WB

Lab

Western blot - Anti-CD45 antibody [EPR20033] (AB208022)

False colour image of Western blot : Anti-CD45 antibody [EPR20033] staining at 1/1000 dilution, shown in green; Mouse anti-Alpha Tubulin [DM1A] (ab7291) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab208022 was shown to bind specifically to CD45. A band was observed at 160-220 kDa in wild-type Jurkat cell lysates with no signal observed at this size in PTPRC CRISPR-Cas9 edited cell line ab274932 (CRISPR-Cas9 edited cell lysate ab274990). The band observed in the CRISPR-Cas9 edited lysate lane below 160-220 kDa is likely to represent a truncated form of CD45. This has not been investigated further and the functional properties of the gene product have not been determined. To generate this image, wild-type and PTPRC CRISPR-Cas9 edited Jurkat cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 5 % milk in TBS-0.1 % Tween^® 20 (TBS-T) 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-CD45 antibody [EPR20033] (ab208022) at 1/1000 dilution

Lane 1:

Wild-type Jurkat cell lysate at 20 µg

Lane 2:

PTPRC knockout Jurkat cell lysate at 20 µg

Lane 3:

THP-1 cell lysate at 20 µg

Lane 4:

MCF7 cell lysate at 20 µg

false

• WB

Supplier Data

Western blot - Anti-CD45 antibody [EPR20033] (AB208022)

Blocking/Dilution buffer : 5% NFDM/TBST.

Exposure time : Lane 1 : 30 seconds; Lane 2 : 5 seconds.

All lanes:

Western blot - Anti-CD45 antibody [EPR20033] (ab208022) at 1/1000 dilution

Lane 1:

J774A.1 (Mouse reticulum cell sarcoma macrophage cell line) whole cell lysate at 20 µg

Lane 2:

WEHI-231 (Mouse B cell lymphoblast cell line) 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 1/100000 dilution

Predicted band size: 147 kDa

Observed band size: 180 kDa

false

• WB

Supplier Data

Western blot - Anti-CD45 antibody [EPR20033] (AB208022)

Blocking/Dilution buffer : 5% NFDM/TBST.

Exposure time : Lane 1 : 3 minutes; Lane 2 : 15 seconds.

All lanes:

Western blot - Anti-CD45 antibody [EPR20033] (ab208022) at 1/1000 dilution

Lane 1:

Mouse thymus lysate at 10 µg

Lane 2:

Mouse spleen tissue lysate at 10 µg

Secondary

Lane 1:

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

Lane 2:

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: 147 kDa

Observed band size: 180 kDa,240 kDa

false