WB
Western blot analysis of SEC61B on different lysates with Rabbit anti-SEC61B antibody at 1/2,000 dilution.
Lane 1: HeLa cell lysate (20 µg/Lane),
Lane 2: HepG2 cell lysate (20 µg/Lane),
Lane 3: Bxpc-3 cell lysate (20 µg/Lane),
Lane 4: C2C12 cell lysate (20 µg/Lane),
Lane 5: C6 cell lysate (20 µg/Lane),
Lane 6: Mouse liver tissue lysate (40 µg/Lane),
Lane 7: Rat liver tissue lysate (40 µg/Lane),
Exposure time: 7 seconds;
4-20% SDS-PAGE gel.
Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody at 1/2,000 dilution was used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody at 1/50,000 dilution was used for 1 hour at room temperature.
IHC
Immunohistochemical analysis of paraffin-embedded human liver tissue with Rabbit anti-SEC61B antibody at 1/200 dilution.
The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 20 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody at 1/200 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.
IHC
Immunohistochemical analysis of paraffin-embedded mouse liver tissue with Rabbit anti-SEC61B antibody at 1/200 dilution.
The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 20 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody at 1/200 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.
IHC
Immunohistochemical analysis of paraffin-embedded rat liver tissue with Rabbit anti-SEC61B antibody at 1/200 dilution.
The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 20 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody at 1/200 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.
IHC
Immunohistochemical analysis of paraffin-embedded mouse cerebellum tissue with Rabbit anti-SEC61B antibody at 1/1,000 dilution.
The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 20 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody at 1/1,000 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.
IHC
Immunohistochemical analysis of paraffin-embedded rat pancreas tissue with Rabbit anti-SEC61B antibody at 1/1,000 dilution.
The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 20 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody at 1/1,000 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.
ICC/IF
Immunocytochemistry analysis of C2C12 cells labeling SEC61B with Rabbit anti-SEC61B antibody at 1/100 dilution.
Cells were fixed in 4% paraformaldehyde for 15 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 15 minutes at room temperature, then blocked with 1% BSA in 10% negative goat serum for 1 hour at room temperature. Cells were then incubated with Rabbit anti-SEC61B antibody at 1/100 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (488) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. Nuclear DNA was labelled in blue with DAPI.
Beta tubulin (red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (594) was used as the secondary antibody at 1/1,000 dilution.
IF-P
Immunofluorescence analysis of paraffin-embedded mouse liver tissue labeling SEC61B with Rabbit anti-SEC61B antibody at 1/200 dilution.
The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 20 minutes. The tissues were blocked in 10% negative goat serum for 1 hour at room temperature, washed with PBS, and then probed with the primary antibody (green) at 1/200 dilution overnight at 4 ℃, washed with PBS. Goat Anti-Rabbit IgG H&L (488) was used as the secondary antibody at 1/1,000 dilution. Nuclei were counterstained with DAPI (blue).
IF-P
Immunofluorescence analysis of paraffin-embedded rat pancreas tissue labeling SEC61B with Rabbit anti-SEC61B antibody at 1/200 dilution.
The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 20 minutes. The tissues were blocked in 10% negative goat serum for 1 hour at room temperature, washed with PBS, and then probed with the primary antibody (green) at 1/200 dilution overnight at 4 ℃, washed with PBS. Goat Anti-Rabbit IgG H&L (488) was used as the secondary antibody at 1/1,000 dilution. Nuclei were counterstained with DAPI (blue).
IP
SEC61B was immunoprecipitated from 0.2 mg HeLa cell lysate with Rabbit anti-SEC61B antibody at 2 µg/10 µl beads. Western blot was performed from the immunoprecipitate using Rabbit anti-SEC61B antibody at 1/2,000 dilution. Mouse Anti-Rabbit IgG kappa light chain secondary antibody at 1/5,000 dilution was used for 1 hour at room temperature.
Lane 1: HeLa cell lysate (input),
Lane 2: Rabbit anti-SEC61B antibody IP in HeLa cell lysate,
Lane 3: Rabbit IgG instead of Rabbit anti-SEC61B antibody in HeLa cell lysate,
Blocking/Dilution buffer: 5% NFDM/TBST,
Exposure time: 2 seconds.