Anti-RYBP antibody [EPR13059(2)] - ChIP Grade

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-RYBP antibody [EPR13059(2)] - ChIP Grade (AB185971)

Immunohistochemical analysis of paraffin-embedded, Human clear cell carcinoma kidney tissue labeling RYBP with ab185971 at a 1/100 dilution. Counter stained with hematoxylin.

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

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-RYBP antibody [EPR13059(2)] - ChIP Grade (AB185971)

Immunohistochemical analysis of paraffin-embedded, Human placenta tissue labeling RYBP with ab185971 at a 1/100 dilution. Counter stained 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-RYBP antibody [EPR13059(2)] - ChIP Grade (AB185971)

Immunofluorescence analysis of, paraformaldehyde-fixed, SW480 cells labeling RYBP with ab185971 at a 1/250 dilution. As secondary antibody goat anti-rabbit IgG (Alexa Fluor®555) was used at a 1/200. In blue DAPI staining.

• Flow Cyt (Intra)

Lab

Flow Cytometry (Intracellular) - Anti-RYBP antibody [EPR13059(2)] - ChIP Grade (AB185971)

Intracellular Flow Cytometry analysis of A431 (Human epidermoid carcinoma epithelial cell) cells labeling RYBP with purified ab185971 at 1/20 dilution (11.7μg/mL) (Red). Cells were fixed with 4% paraformaldehyde and permeabilised with 90% methanol. A Goat anti rabbit IgG (Alexa Fluor^® 488, ab150077) secondary antibody was used at 1/2000 dilution. Isotype control - Rabbit monoclonal IgG (ab172730) (black). Unlabeled control - Unlabelled cells (blue).

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-RYBP antibody [EPR13059(2)] - ChIP Grade (AB185971)

Immunofluorescence analysis of, paraformaldehyde-fixed, HepG2 cells labeling RYBP with ab185971 at a 1/250 dilution. As secondary antibody goat anti-rabbit IgG (Alexa Fluor®555) was used at a 1/200. In blue DAPI staining.

• Flow Cyt (Intra)

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Flow Cytometry (Intracellular) - Anti-RYBP antibody [EPR13059(2)] - ChIP Grade (AB185971)

Intracellular Flow Cytometry analysis of paraformaldehyde-fixed K562 cells labeling RYBP with ab185971 at a 1/40 dilution and secondary antibody goat anti-rabbit IgG (FITC, red) at a 1/150 dilution, or negative control rabbit IgG (green).

• IP

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Immunoprecipitation - Anti-RYBP antibody [EPR13059(2)] - ChIP Grade (AB185971)

Western blot analysis on immunoprecipitation pellet from HepG2 cell lysate labeling RYBP with ab185971 at 1/50 dilution and goat anti-rabbit IgG, (H+L), peroxidase conjugated at a 1/1000 dilution.

All lanes:

Immunoprecipitation - Anti-RYBP antibody [EPR13059(2)] - ChIP Grade (ab185971)

Predicted band size: 25 kDa

Observed band size: 32 kDa

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• ChIP

Unknown

ChIP - Anti-RYBP antibody [EPR13059(2)] - ChIP Grade (AB185971)

Chromatin was prepared from HEK293 cells according to the Abcam Dual X-ChIP protocol*. Cells were fixed with EGS for 30 minutes, then formaldehyde for 10 minutes.
The ChIP was performed with 25 µg of chromatin, 5 µg of ab185971 (red), and 20 µl of Protein A/G sepharose beads. 5 µg of rabbit normal IgG was added to the beads control (gray). The immunoprecipitated DNA was quantified by real time PCR (Sybr green approach).
Primers and probes are located in the first kb of the transcribed region.
*http : //www.abcam.com/resources?keywords=X%20ChIP%20protocol

• ChIP

Unknown

ChIP - Anti-RYBP antibody [EPR13059(2)] - ChIP Grade (AB185971)

Chromatin was prepared from NIH/3T3 cells according to the Abcam Dual X-ChIP protocol*. Cells were fixed with EGS for 30 minutes, then formaldehyde for 10 minutes.
The ChIP was performed with 25 µg of chromatin, 5 µg of ab185971 (red), and 20 µl of Protein A/G sepharose beads. 5 µg of rabbit normal IgG was added to the beads control (gray). The immunoprecipitated DNA was quantified by real time PCR (Sybr green approach).
Primers and probes are located in the first kb of the transcribed region.
*http : //www.abcam.com/resources?keywords=X%20ChIP%20protocol

• IP

Lab

Immunoprecipitation - Anti-RYBP antibody [EPR13059(2)] - ChIP Grade (AB185971)

ab185971 (purified) at 1/20 dilution (11.7μg/mL) immunoprecipitating RYBP in NIH/3T3 (Mouse embryonic fibroblast) whole cell lysate 10 μg.

Lane 1 (input) : NIH/3T3 (Mouse embryonic fibroblast) whole cell lysate 10 μg
Lane 2 (+) : ab185971 & NIH/3T3 whole cell lysate
Lane 3 (-) : Rabbit monoclonal IgG (ab172730) instead of ab185971 in NIH/3T3 whole cell lysate

For western blotting, ab185971 at 1/500 dilution (0.468 μg/mL) and veriBlot for IP secondary antibody (HRP) (ab131366) at 1/1000 dilution was used.

Blocking and diluting buffer : 5% NFDM /TBST.

All lanes:

Immunoprecipitation - Anti-RYBP antibody [EPR13059(2)] - ChIP Grade (ab185971)

Predicted band size: 25 kDa

false

• WB

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Western blot - Anti-RYBP antibody [EPR13059(2)] - ChIP Grade (AB185971)

All lanes:

Western blot - Anti-RYBP antibody [EPR13059(2)] - ChIP Grade (ab185971) at 1/10000 dilution

All lanes:

HepG2 cell lysate at 20 µg

Secondary

All lanes:

goat anti-rabbit IgG, (H+L), peroxidase conjugated at 1/1000 dilution

Predicted band size: 25 kDa

Observed band size: 32 kDa

true

• WB

Supplier Data

Western blot - Anti-RYBP antibody [EPR13059(2)] - ChIP Grade (AB185971)

All lanes:

Western blot - Anti-RYBP antibody [EPR13059(2)] - ChIP Grade (ab185971) at 1/1000 dilution

Lane 1:

K562 cell lysate at 20 µg

Lane 2:

SW480 cell lysate at 20 µg

Secondary

All lanes:

goat anti-rabbit IgG, (H+L), peroxidase conjugated at 1/1000 dilution

Predicted band size: 25 kDa

Observed band size: 32 kDa

true

• WB

Supplier Data

Western blot - Anti-RYBP antibody [EPR13059(2)] - ChIP Grade (AB185971)

All lanes:

Western blot - Anti-RYBP antibody [EPR13059(2)] - ChIP Grade (ab185971) at 1/1000 dilution

All lanes:

rat PC12 cell lysate at 10 µg

Secondary

All lanes:

goat anti-rabbit IgG, (H+L), peroxidase conjugated at 1/1000 dilution

Predicted band size: 25 kDa

Observed band size: 32 kDa

true