Anti-SEC23B 抗体 [EPR22723-24] (ab245212)

SEC23B was immunoprecipitated from 0.35 mg MEF (mouse embryonic fibroblast (immortalized)) whole cell lysate 20 ug with ab245212 at 1/30. Western blot was performed on the immunoprecipitate using ab245212 at 1/500 dilution. VeriBlot for IP Detection Reagent (HRP)(ab131366) was used at 1/1000 dilution.

Lane 1: MEF whole cell lysate 20 ug.

Lane 2: ab245212 IP in MEF whole cell lysate.

Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab245212 in MEF whole cell lysate.

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

Exposure time: 1 second.

Intracellular flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized MEF (Mouse embryonic fibroblast (immortalized)) cells labeling SEC23B with ab245212 at 1/500 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. 

ab245212 staining SEC23B in Raw264.7 cells, with negative expression in cells. The cells were fixed with 4% formaldehyde (10 min), permeabilised with 0.1% Triton x-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated overnight at +4°C with ab245212 at 1 µg/ml and ab7291, Mouse monoclonal [DM1A] to alpha Tubulin at 0.5 µg/ml. Cells were then incubated with ab150081, Goat polyclonal Secondary Antibody to Rabbit IgG - H&L (Alexa Fluor^® 488), pre-adsorbed at 1/1000 dilution (shown in green) and ab150119, Goat polyclonal Secondary Antibody to Mouse IgG - H&L (Alexa Fluor^® 647), pre-adsorbed at 1/1000 dilution (shown in red). Nuclear DNA was labelled with DAPI (shown in blue).

Image was acquired with a confocal microscope (Leica-Microsystems TCS SP8) and a single confocal section is shown.

Immunohistochemical analysis of paraffin-embedded mouse liver tissue labeling SEC23B with 245212 at 1/1/1000 followed by a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Positive staining on mouse liver is observed (PMID: 22745161).
The section was incubated with ab245212 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.

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

The molecular weight observed is consistent with what has been described in the literature (PMID: 22745161).

Exposure times: Lane 1: 26 seconds; Lane 2: 3 seconds; Lanes 3-4: 6 seconds.

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized MEF cells labeling SEC23B with ab245212 at 1/50 dilution, followed by ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) antibody at 1/1000 dilution (Green). Confocol image showing mainly cytoplasmic staining in MEFs. ab195889 Anti-alpha Tubulin antibody (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.

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

The lysates were kindly provided by an anonymous collaborator.  Exposure time: 59 seconds.

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

Exposure time: 26 seconds.