重组Anti-CSL4抗体[EPR13525]

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CSL4 antibody [EPR13525] (AB181167)

Immunohistochemical analysis of paraffin embedded Human breast carcinoma tissue labeling CSL4 with ab181167 at a 1/100 dilution. Prediluted HRP Polymer for Rabbit IgG secondary used. Counterstained with Hematoxylin.

Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-CSL4 antibody [EPR13525] (AB181167)

Immunofluorescence analysis of HepG2 cells (fixative 4% paraformaldehyde) labeling CSL4 with ab181167 at a 1/250 dilution. Goat anti rabbit IgG (Alexa Fluor® 555) secondary used at a 1/200 diution.

• Flow Cyt (Intra)

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Flow Cytometry (Intracellular) - Anti-CSL4 antibody [EPR13525] (AB181167)

Intracellular Flow Cytometry analysis of K562 cells using ab181167at a 1/90 dilution (red) or a Rabbit monoclonal IgG (negative) (green). Goat anti rabbit IgG (FITC) secondary used at a 1/150 dilution.

• IP

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Immunoprecipitation - Anti-CSL4 antibody [EPR13525] (AB181167)

Detection of Human CSL4 by Western Blot of Immunoprecipitate. 293T whole cell lysate (lane 1) and a negative control (lane 2) loaded, ab181167 used for blotting immunoprecipitated CSL4 at a 1/50 dilution. Anti-Rabbit IgG (HRP), specific to the non-reduced form of IgG secondary used at a 1/1500 dilution.

All lanes:

Immunoprecipitation - Anti-CSL4 antibody [EPR13525] (ab181167)

Predicted band size: 21 kDa

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• WB

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Western blot - Anti-CSL4 antibody [EPR13525] (AB181167)

All lanes:

Western blot - Anti-CSL4 antibody [EPR13525] (ab181167) at 1/2000 dilution

Lane 1:

293T cell lysate at 20 µg

Lane 2:

Jurkat cell lysate at 20 µg

Lane 3:

HepG2 cell lysate at 20 µg

Secondary

All lanes:

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

Predicted band size: 21 kDa

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• WB

Lab

Western blot - Anti-CSL4 antibody [EPR13525] (AB181167)

False colour image of Western blot : Anti-CSL4 antibody [EPR13525] 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, ab181167 was shown to bind specifically to CSL4. A band was observed at 22 kDa in wild-type HeLa cell lysates with no signal observed at this size in EXOSC1 CRISPR-Cas9 edited cell line ab265194 (EXOSC1 CRISPR-Cas9 edited cell lysate ab257945). The band observed in the CRISPR-Cas9 edited lysate lane above 22 kDa is likely to represent CSL4 with an insertion. 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 EXOSC1 CRISPR-Cas9 edited HeLa cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3% 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-CSL4 antibody [EPR13525] (ab181167) at 1/1000 dilution

Lane 1:

Wild-type HeLa cell lysate at 20 µg

Lane 2:

EXOSC1 CRISPR-Cas9 edited HeLa cell lysate at 20 µg

Lane 3:

PC-12 cell lysate at 20 µg

Lane 4:

SH-SY5Y cell lysate at 20 µg

Predicted band size: 21 kDa

Observed band size: 22 kDa

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