All lanes : Anti-GC1q R antibody [EPR23238-6] (ab270033) at 1/1000 dilution

Lane 1 : HeLa (human cervix adenocarcinoma epithelial cell) whole cell lysate
Lane 2 : A549 (human lung carcinoma epithelial cell) whole cell lysate
Lane 3 : MCF7 (human breast adenocarcinoma epithelial cell) whole cell lysate
Lane 4 : WI-38 ( human fetal lung fibroblast) 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: 31 kDa

The expression profile/ molecular weight observed is consistent with what has been described in the literature (PMID: 20100866).

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

Exposure time: Lanes 1-3: 15 secs; Lane 4: 3 secs.

All lanes : Anti-GC1q R antibody [EPR23238-6] (ab270033) at 1/1000 dilution

Lane 1 : Human lung cancer tissue lysate
Lane 2 : Human spleen tissue lysate

Lysates/proteins at 20 µg per lane.

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

Predicted band size: 31 kDa

The expression profile/ molecular weight observed is consistent with what has been described in the literature (PMID: 20100866).

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

Exposure time: 15 secs.

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized A549 cells labeling GC1q R with ab270033 at 1/500 dilution, followed by ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) antibody at 1/1000 dilution (Green). Confocal image showing mitochondrial staining in A549 cell line. ab33985 Anti-COX IV antibody (human) was used to counterstain tubulin at 1/1000 dilution (Red). The nuclear counterstain was DAPI (Blue).

-ve control 1: Stained with ab270033 at 1/500 followed by ab150120 Goat Anti-Mouse IgG H&L (Alexa Fluor^® 594) at a 1/1000 dilution.

-ve control 2: Stained with ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) at 1/1000 dilution followed by ab33985 Anti-COX IV antibody (human) at 1/1000 dilution.

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HeLa cells labeling GC1q R with ab270033 at 1/500 dilution, followed by ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) antibody at 1/1000 dilution (Green). Confocal image showing mitochondrial staining in HeLa cell line. ab33985 Anti-COX IV antibody (human) was used to counterstain tubulin at 1/1000 dilution (Red). The nuclear counterstain was DAPI (Blue).

-ve control 1: Stained with ab270033 at 1/500 followed by ab150120 Goat Anti-Mouse IgG H&L (Alexa Fluor^® 594) at a 1/1000 diliution.

-ve control 2: Stained with ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) at 1/1000 dilution followed by ab33985 Anti-COX IV antibody (human) at 1/1000 dilution.

Flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized HeLa (Human cervix adenocarcinoma epithelial cell) cells labeling GC1q R with ab270033 at 1/600 dilution (0.1µg) (Red) compared with a Rabbit monoclonal IgG (ab172730) (Black) isotype control and an unlabeled 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 cerebral cortex tissue labeling GC1q R with ab270033 at 1/500 (1.27 μg/ml) followed by a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Positive staining on human cerebral cortex is observed (PMID: 23924515). The section was incubated with ab270033 for 30 mins at room temperature.The immunostaining was performed on a Leica Biosystems BOND^® RX instrument Counterstained with hematoxylin. Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins.

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).