Immunohistochemical analysis of paraffin-embedded mouse testis tissue stained for YTHDC1 with ab220159 at 1/500 dilution, followed by a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Nuclear staining on mouse testis (PMID: 27602518; PMID: 29103884) is observed. Counterstained with Hematoxylin.

Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). 

The section was incubated with ab220159 for 30 mins at 37℃. The immunostaining staining was performed on a Leica Biosystems BOND^® RX instrument.

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

Immunohistochemical analysis of paraffin-embedded rat spleen tissue stained for YTHDC1 with ab220159 at 1/500 dilution, followed by a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Nuclear staining on rat spleen (PMID: 27602518; PMID: 29103884) is observed. Counterstained with Hematoxylin.

Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). 

The section was incubated with ab220159 for 30 mins at 37℃. The immunostaining staining was performed on a Leica Biosystems BOND^® RX instrument.

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

Immunohistochemical analysis of paraffin-embedded human endometrial carcinoma tissue stained for YTHDC1 with ab220159 at 1/500 dilution, followed by a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Nuclear staining on human endometrial carcinoma (PMID: 27602518; PMID: 29103884) is observed. Counterstained with Hematoxylin.

Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). 

The section was incubated with ab220159 for 30 mins at 37℃. The immunostaining staining was performed on a Leica Biosystems BOND^® RX instrument.

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

Immunohistochemical analysis of paraffin-embedded human testis tissue stained for YTHDC1 with ab220159 at 1/500 dilution, followed by a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Nuclear staining on human testis (PMID: 27602518; PMID: 29103884) is observed. Counterstained with Hematoxylin.

Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). 

The section was incubated with ab220159 for 30 mins at 37℃. The immunostaining staining was performed on a Leica Biosystems BOND^® RX instrument.

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

All lanes : Anti-YTHDC1 antibody [EPR21821] (ab220159) at 1/5000 dilution

Lane 1 : HeLa (human cervix adenocarcinoma epithelial cell) transfected with scrambled siRNA control whole cell lysate
Lane 2 : YTHDC1 knockdown HeLa whole cell lysate

Lysates/proteins at 20 µg per lane.

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

Predicted band size: 85 kDa
Observed band size: 100 kDa why is the actual band size different from the predicted?


Exposure time: 59 seconds

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

The control and YTHDC1 knockdown cell lysates were kindly provided by an anonymous collaborator.

Lane 1 : Anti-YTHDC1 antibody [EPR21821] (ab220159) at 1/5000 dilution
Lanes 2-5 : Anti-YTHDC1 antibody [EPR21821] (ab220159) at 1/1000 dilution

Lane 1 : K-562 (human chronic myelogenous leukemia lymphoblast) whole cell lysate
Lane 2 : C6 (rat glial tumor glial cell) whole cell lysate
Lane 3 : RAW 264.7 (mouse abelson murine leukemia virus-induced tumor macrophage) whole cell lysate
Lane 4 : PC-12 (rat adrenal gland pheochromocytoma) whole cell lysate
Lane 5 : NIH/3T3 (mouse embryonic fibroblast) whole cell lysate

Lysates/proteins at 10 µg per lane.

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

Predicted band size: 85 kDa
Observed band size: 100,95 kDa why is the actual band size different from the predicted?

Blocking/diluting buffer and concentration: 5% NFDM/TBST

The doublet observed is consistent with what has been shown in the literature (PMID: 26876937). The second band may represent the isoform 2 of YTHDC1 (PMID: 23765422).

Exposure times:

Lane 1:70 seconds;

Lanes 2-5: 48 seconds.

Anti-YTHDC1 antibody [EPR21821] (ab220159) at 1/1000 dilution + Human brain tissue lysate at 20 µg

Secondary
VeriBlot for IP Detection Reagent (HRP) (ab131366) at 1/1000 dilution

Predicted band size: 85 kDa
Observed band size: 100,95 kDa why is the actual band size different from the predicted?


Exposure time: 3 minutes

Blocking/Diluting buffer and concentration: 5% NFDM/TBST

The doublet observed is consistent with what has been shown in the literature (PMID: 26876937). The second band may represent the isoform 2 of YTHDC1 (PMID: 23765422).