All lanes : Anti-Chromogranin C/SGII antibody [EPR23876-52] (ab275019) at 1/1000 dilution

Lane 1 : Mouse hypothalamus tissue lysate
Lane 2 : Rat hypothalamus tissue lysate

Lysates/proteins at 40 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated (ab97051) at 1/20000 dilution

Predicted band size: 71 kDa

Blocking and dilution buffer:  5% NFDM/TBST.

Exposure times: Lane 1: 26 seconds; Lane 2: 48 seconds.

Chromogranin C/SGII is proteolytically processed to small fragments. 81-kDa, 22 to 48 kDa forms are observed. The molecular weight observed is consistent with what has been described in the literature (PMID: 2282535, 8492910, 16357044, 7883050).

Immunofluorescent analysis of 100% methanol-fixed PC-12 cells labelling Chromogranin C/SGII with ab275019 at 1/50 (10.68 ug/ml) dilution, followed by ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) antibody at 1/1000 dilution (Green). Confocal image showing cytoplasmic staining in PC-12 cell line. is observed. ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor^® 594) was used to counterstain tubulin at 1/200 dilution (Red). The Nuclear counterstain was DAPI (Blue).

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

Immunohistochemical analysis of paraffin-embedded Rat pancreas tissue labeling Chromogranin C/SGII with ab275019 at 1/2000 dilution followed by a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Cytoplasmic staining in pancreatic islet (PMID: 30690892). The section was incubated with ab275019 for 10 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND^® RX instrument.Counterstained with Hematoxylin.

Secondary antibody only control: Secondary antibody is a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101).

Heat mediated antigen retrieval with Citrate buffer (pH 6.0, epitope retrieval solution 1) for 20 mins.

All lanes : Anti-Chromogranin C/SGII antibody [EPR23876-52] (ab275019) at 1/1000 dilution

Lane 1 : SH-SY5Y (human neuroblastoma epithelial cell) whole cell lysate
Lane 2 : Neuro-2a (mouse neuroblastoma neuroblast) whole cell lysate
Lane 3 : Beta-TC-6 (mouse pancreas insulinoma beta cell) whole cell lysate
Lane 4 : GH3 (rat pituitary epithelial) whole cell lysate
Lane 5 : PC-12 (rat adrenal gland pheochromocytoma) whole cell lysate

Lysates/proteins at 40 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated (ab97051) at 1/20000 dilution

Predicted band size: 71 kDa

Blocking and dilution buffer: 5% NFDM/TBST.

Exposure times: Lane 1: 37 seconds; Lane 2: 15 seconds; Lane3-4: 5.5 seconds; Lane5: 1 second.

Chromogranin C/SGII is proteolytically processed to small fragments. 81-kDa, 55-kDa, 22 to 48 kDa forms are observed. The molecular weight observed is consistent with what has been described in the literature (PMID: 2282535, 8492910, 16357044, 7883050).

 

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized Beta-TC-6 cells labelling Chromogranin C/SGII with ab275019 at 1/50 (10.68 ug/ml) dilution, followed by ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) antibody at 1/1000 dilution (Green). Confocal image showing cytoplasmic staining in Beta-TC-6 cell line. ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor^® 594) was used to counterstain tubulin at 1/200 dilution (Red). The Nuclear counterstain was DAPI (Blue).

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

Immunohistochemical analysis of paraffin-embedded Mouse pancreas tissue labeling Chromogranin C/SGII with ab275019 at 1/2000 dilution followed by a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Cytoplasmic staining in mouse pancreatic islet (PMID: 30690892). The section was incubated with ab275019 for 10 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND^® RX instrument.Counterstained with Hematoxylin.

Secondary antibody only control: Secondary antibody is a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101).

Heat mediated antigen retrieval with Citrate buffer (pH 6.0, epitope retrieval solution 1) for 20 mins.

Immunohistochemical analysis of 4% PFA-fixed, 0.2% Triton X-100 permeabilized frozen Rat pancreas tissue labeling Chromogranin C/SGII with ab275019 at 1/500 (1.068 ug/ml) dilution followed by ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) at 1/1000 dilution (Green). Positive staining on rat pancreatic islet is observed. The nuclear counterstain was DAPI (Blue). Insulin was counterstained using ab6995 at 1/1000 dilution, followed by ab150120 Goat Anti-Mouse IgG H&L (Alexa Fluor^® 594) at 1/1000 dilution (Red).

Secondary antibody control: Secondary antibody is ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488)at 1/1000 dilution.

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

Flow cytometric analysis of 4% paraformaldehyde fixed, 90% methanol permeabilized PC-12 (Rat adrenal gland pheochromocytoma) cells labelling Chromogranin C/SGII with ab275019 at 1/500 dilution (0.1ug) (Red) compared with a Rabbit monoclonal IgG (ab172730) isotype control (Black)  and an unlabelled control (cells without incubation with primary antibody and secondary antibody) (Blue).

A Goat anti rabbit IgG (Alexa Fluor^® 488, ab150077) at 1/2000 dilution was used as the secondary antibody.

Immunohistochemical analysis of 4% PFA-fixed, 0.2% Triton X-100 permeabilized frozen Mouse pancreas tissue labeling Chromogranin C/SGII with ab275019 at 1/500 (1.068 ug/ml) dilution followed by ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) at 1/1000 dilution (Green). Positive staining on mouse pancreatic islet is observed. Insulin was counterstained using ab6995 at 1/1000 dilution, followed by ab150120 Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) at 1/1000 dilution (Red).The nuclear counterstain was DAPI (Blue).

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

Flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized Beta-TC-6 (Mouse pancreas insulinoma beta cell) cells labelling Chromogranin C/SGII with ab275019 at 1/500 dilution (0.1ug) (Red) compared with a Rabbit monoclonal IgG (ab172730) isotype control (Black) and an unlabelled control (cells without incubation with primary antibody and secondary antibody) (Blue).

A Goat anti rabbit IgG (Alexa Fluor^® 488, ab150077) at 1/2000 dilution was used as the secondary antibody.

Immunohistochemical analysis of paraffin-embedded Human pancreas tissue labeling Chromogranin C/SGII with ab275019 at 1/2000 (0.267 ug/ml) followed by a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Cytoplasmic staining in human pancreatic islet (PMID: 30690892)The section was incubated with ab275019 for 30 mins at room temperature.The immunostaining was performed on a Leica Biosystems BOND® RX instrument Counterstained with Hematoxylin.

Secondary antibody only control: Secondary antibody is a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101).

Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins