Immunocytochemistry analysis of mESCs cultured in 2i conditions labelling DAZL with ab215718 at 1/500 dilution. Cells were fixed with 4% formaldehyde and permeabilized with 0.1% Triton X-100. Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) at 1/4000 was used as the secondary antibody (green). Nuclear DNA was labelled with DAPI (blue).

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Immunohistochemical analysis of paraffin-embedded human testis tissue labeling DAZL with ab215718 at 1/8000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use. Cytoplasmic staining in primary spermatocytes of human testis (PMID: 24746554) is observed. Counterstained with hematoxylin.

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

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

Immunohistochemical analysis of 4% paraformaldehyde-fixed, 0.2% Triton-X-100 permeabilized frozen mouse testis tissue labeling DAZL with ab215718 at 1/300 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor^® staining in the pachytene spermatocytes and secondary spermatocytes, not present in Sertoli, Leydig or myoid cells (PMID: 24746554).

The nuclear counter stain is DAPI (blue).

Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution.

All lanes : Anti-DAZL antibody [EPR21028] (ab215718) at 1/1000 dilution

Lane 1 : Mouse testis tissue lysate
Lane 2 : DAZL knockout mouse testis tissue lysate

Lysates/proteins at 25 µg per lane.

Secondary
All lanes : HRP-conjugated horse anti-rabbit IgG at 1/10000 dilution

Developed using the ECL technique.

Performed under non-reducing conditions.

Predicted band size: 33 kDa
Observed band size: 37 kDa why is the actual band size different from the predicted?


Exposure time: 3 seconds

10% gel.

Blocked with 5% milk for 1 hour at room temperature.

Incubated with the primary antibody for 12 hours at 4°C in TBST.

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All lanes : Anti-DAZL antibody [EPR21028] (ab215718) at 1/1000 dilution

Lane 1 : Mouse testis lysate
Lane 2 : Rat testis lysate
Lane 3 : Human testis lysate

Lysates/proteins at 20 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

Developed using the ECL technique.

Predicted band size: 33 kDa
Observed band size: 38 kDa why is the actual band size different from the predicted?


Exposure time: 3 minutes

Blocking/Dilution buffer: 5% NFDM/TBST.

The blot was developed on a BIO-RAD^® ChemiDoc™ MP instrument.

DAZL was immunoprecipitated from 0.35 mg mouse testis lysate with ab215718 at 1/30 dilution. Western blot was performed from the immunoprecipitate using ab215718 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/10,000 dilution.

Lane 1: Mouse testis lysate 10 μg (Input).

Lane 2: ab215718 IP in mouse testis lysate.

Lane 3: Rabbit monoclonal IgG (ab172730) instead of  ab215718 in mouse testis lysate.

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

Exposure time: 3 seconds.

Immunohistochemical analysis of paraffin-embedded mouse testis tissue labeling DAZL with ab215718 at 1/4000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use. Cytoplasmic staining in spermatogonia and primary spermatocytes of mouse testis (PMID: 24746554) is observed. Counterstained with hematoxylin.

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

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

Immunohistochemical analysis of paraffin-embedded rat testis tissue labeling DAZL with ab215718 at 1/4000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use. Cytoplasmic staining in spermatogonia and primary spermatocytes of rat testis (PMID: 24746554) is observed. Counterstained with hematoxylin

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

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

Immunohistochemical analysis of 4% paraformaldehyde-fixed, 0.2% Triton-X-100 permeabilized frozen rat testis tissue labeling DAZL with ab215718 at 1/60 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Cytoplasmic staining in the pachytene spermatocytes and secondary spermatocytes, not present in Sertoli, Leydig or myoid cells (PMID: 24746554).

The nuclear counter stain is DAPI (blue).

Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution.