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.

 

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.

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.

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.

All lanes : Anti-SEC23B antibody [EPR22723-24] (ab245212) at 1/1000 dilution

Lane 1 : MEF (mouse embryonic fibroblast (immortalized)) whole cell lysate
Lane 2 : Mouse liver tissue lysate
Lane 3 : RAW 264.7 (mouse abelson murine leukemia virus-induced tumor macrophage) whole cell lysate
Lane 4 : NIH/3T3 (mouse embryonic 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: 86 kDa

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.

All lanes : Anti-SEC23B antibody [EPR22723-24] (ab245212) at 1/1000 dilution

Lane 1 : MEF (mouse embryonic fibroblast (immortalized)) transfected with scrambled siRNA control whole cell lysate
Lane 2 : MEF (mouse embryonic fibroblast (immortalized)) transfected with siRNA specifically targeting SEC23B 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: 86 kDa

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

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

All lanes : Anti-SEC23B antibody [EPR22723-24] (ab245212) at 1/5000 dilution

Lane 1 : Purified LIF/His-tagged SEC23B recombinant protein, 10 ng
Lane 2 : His-tagged SEC23A recombinant protein, 10 ng

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

Predicted band size: 86 kDa

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

Exposure time: 26 seconds.