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Western blot analysis of Beta Catenin on different lysates with Mouse anti-Beta Catenin antibody (HA601179) at 1/1,000 dilution.
Lane 1: 293T cell lysate
Lane 2: A431 cell lysate
Lane 3: NCCIT cell lysate
Lane 4: SW480 cell lysate
Lane 5: HT-29 cell lysate
Lane 6: HCT 116 cell lysate
Lysates/proteins at 20 µg/Lane.
Predicted band size: 85 kDa
Observed band size: 85 kDa
Exposure time: 5 minutes;
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 (HA601179) at 1/1,000 dilution was used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Mouse IgG - HRP Secondary Antibody (HA1006) at 1/50,000 dilution was used for 1 hour at room temperature.
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Immunocytochemistry analysis of MCF7 cells labeling Beta Catenin with Mouse anti-Beta Catenin antibody (HA601179) at 1/100 dilution.
Cells were fixed in 4% paraformaldehyde for 20 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 5 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 Mouse anti-Beta Catenin antibody (HA601179) at 1/100 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Mouse IgG H&L (iFluor™ 488, HA1125) 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 (ET1602-4, red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Rabbit IgG H&L (iFluor™ 594, HA1122) were used as the secondary antibody at 1/1,000 dilution.
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Flow cytometric analysis of MCF7 cells labeling Beta Catenin.
Cells were fixed and permeabilized. Then stained with the primary antibody (HA601179, 1μg/mL) (red) compared with Mouse IgG1 Isotype Control (green). After incubation of the primary antibody at +4℃ for an hour, the cells were stained with a iFluor™ 594 conjugate-Goat anti-Mouse IgG Secondary antibody (HA1126) at 1/1,000 dilution for 30 minutes at +4℃. Unlabelled sample was used as a control (cells without incubation with primary antibody; black).
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