Anti-YTHDF3 antibody [EPR21912-3] - BSA and Azide free

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-YTHDF3 antibody [EPR21912-3] - BSA and Azide free (AB255267)

Immunohistochemical analysis of paraffin-embedded human colon tissue labeling YTHDF3 with ab220161 at 1/100 dilution, followed by a ready to use Goat Anti-Rabbit IgG H&L (HRP). Cytoplasmic staining on human colon (PMID : 29103884, 28250115) is observed. Counterstained with hematoxylin.

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is a ready to use Goat Anti-Rabbit IgG H&L (HRP).

Perform heat mediated antigen retrieval using ab93684 (Tris/EDTA buffer, pH 9.0).

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab220161).

• Flow Cyt (Intra)

Unknown

Flow Cytometry (Intracellular) - Anti-YTHDF3 antibody [EPR21912-3] - BSA and Azide free (AB255267)

Intracellular flow cytometric analysis of 4% paraformaldehyde-fixed, 90% methanol permeabilized HeLa (human cervix adenocarcinoma epithelial cell) cell line labeling YTHDF3 with ab220161 at 1/60 (red) compared with a Rabbit monoclonal IgG (ab172730) (black) and an unlabeled control (cells without incubation with primary antibody and secondary antibody) (blue). Goat anti rabbit IgG (Alexa Fluor^® 488, ab150077), at 1/2000 dilution 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 (ab220161).

• IP

Unknown

Immunoprecipitation - Anti-YTHDF3 antibody [EPR21912-3] - BSA and Azide free (AB255267)

YTHDF3 was immunoprecipitated from 0.35 mg HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysate with ab220161 at 1/30 dilution. Western blot was performed from the immunoprecipitate using ab220161 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used at 1/5000 dilution.

Lane 1 : HeLa whole cell lysate 10 μg (Input).
Lane 2 : ab220161 IP in HeLa whole cell lysate.
Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab220161 in HeLa whole cell lysate.

Blocking/Dilution buffer : 5% NFDM/TBST.
Exposure time : 15 seconds.

Lysate were made freshly and used in IP test immediately to minimize protein degradation. Incubation time was 2h.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab220161).

All lanes:

Immunoprecipitation - Anti-YTHDF3 antibody [EPR21912-3] (<a href='/en-us/products/primary-antibodies/ythdf3-antibody-epr21912-3-ab220161'>ab220161</a>)

Predicted band size: 64 kDa

Observed band size: 73 kDa

false

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-YTHDF3 antibody [EPR21912-3] - BSA and Azide free (AB255267)

Immunohistochemical analysis of paraffin-embedded rat cerebrum tissue labeling YTHDF3 with ab220161 at 1/500 dilution, followed by a ready to use Goat Anti-Rabbit IgG H&L (HRP). Cytoplasmic staining on neurons of rat cerebrum (PMID : 29103884, 28250115) is observed. Counterstained with hematoxylin.

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is a ready to use Goat Anti-Rabbit IgG H&L (HRP).

Perform heat mediated antigen retrieval using ab93684 (Tris/EDTA buffer, pH 9.0).

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab220161).

• IHC-Fr

Unknown

Immunohistochemistry (Frozen sections) - Anti-YTHDF3 antibody [EPR21912-3] - BSA and Azide free (AB255267)

Immunohistochemical analysis of 4% PFA-fixed, 0.2% Triton X-100 permeabilized frozen rat cerebrum tissue labeling YTHDF3 with ab220161 at 1/100 dilution (green), followed by ab150077 AlexaFluor^®488 Goat anti-Rabbit secondary at a 1/1,000 dilution. Cytoplasmic and nuclear staining in rat cerebrum (PMID : 29103884, 28250115) is observed. Counterstained with DAPI (blue).

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is ab150077 AlexaFluor^®488 Goat anti-Rabbit used at a 1/1,000 dilution.

Heat mediated antigen retrieval using sodium citrate buffer (10 mM citrate pH 6.0 and 0.05% Tween-20).

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab220161).

• IHC-Fr

Unknown

Immunohistochemistry (Frozen sections) - Anti-YTHDF3 antibody [EPR21912-3] - BSA and Azide free (AB255267)

Immunohistochemical analysis of 4% PFA-fixed, 0.2% Triton X-100 permeabilized frozen mouse cerebrum tissue labeling YTHDF3 with ab220161 at 1/100 dilution (green), followed by ab150077 AlexaFluor^®488 Goat anti-Rabbit secondary at a 1/1,000 dilution. Cytoplasmic and nuclear staining in mouse cerebrum (PMID : 29103884, 28250115) is observed. Counterstained with DAPI (blue).

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is ab150077 AlexaFluor^®488 Goat anti-Rabbit used at a 1/1,000 dilution.

Heat mediated antigen retrieval using sodium citrate buffer (10 mM citrate pH 6.0 and 0.05% Tween-20).

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab220161).

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-YTHDF3 antibody [EPR21912-3] - BSA and Azide free (AB255267)

Immunohistochemical analysis of paraffin-embedded mouse cerebrum tissue labeling YTHDF3 with ab220161 at 1/500 dilution, followed by a ready to use Goat Anti-Rabbit IgG H&L (HRP). Cytoplasmic staining on neurons of mouse cerebrum (PMID : 29103884, 28250115) is observed. Counterstained with hematoxylin.

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is a ready to use Goat Anti-Rabbit IgG H&L (HRP).

Perform heat mediated antigen retrieval using ab93684 (Tris/EDTA buffer, pH 9.0).

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab220161).

• WB

Lab

Western blot - Anti-YTHDF3 antibody [EPR21912-3] - BSA and Azide free (AB255267)

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab220161).

Western blot : Anti-YTHDF3 antibody [EPR21912-3] ab220161 staining at 1/1000 dilution, shown in green; Mouse anti GAPDH ab8245 loading control staining at 1/20000 dilution, shown in magenta. A band was observed at 70 kDa in Wild-type PC-3 cell lysates with no signal observed at this size in YTHDF3 knockout PC-3 cell line. To generate this image, 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 800CW and Goat anti-Mouse 680RD at 1/20,000 dilution.

All lanes:

Western blot - Anti-YTHDF3 antibody [EPR21912-3] (<a href='/en-us/products/primary-antibodies/ythdf3-antibody-epr21912-3-ab220161'>ab220161</a>) at 1/1000 dilution

Lane 1:

Wild-type PC-3 cell lysates at 20 µg

Lane 2:

Western blot - Human YTHDF3 knockout PC-3 cell line (<a href='/en-us/products/cell-lines/human-ythdf3-knockout-pc-3-cell-line-ab290587'>ab290587</a>) at 20 µg

Lane 3:

HeLa cell lysates at 20 µg

Lane 4:

HT-1080 cell lysates at 20 µg

Secondary

All lanes:

Goat anti-Rabbit 800CW & Goat anti-Mouse 680RD at 1/20000 dilution

Predicted band size: 64 kDa

Observed band size: 70 kDa

false