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☑ Knockdown (KD)
Western blot analysis of Staufen on different lysates with Rabbit anti-Staufen antibody (ET1706-41) at 1/1,000 dilution.
Lane 1: HAP1-parental cell lysate
Lane 2: HAP1-Staufen KD cell lysate
Lysates/proteins at 10 µg/Lane.
Predicted band size: 63 kDa
Observed band size: 63/55 kDa
Exposure time: 1 minute; ECL: K1801;
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 (ET1706-41) at 1/1,000 dilution was used in primary antibody dilution (K1803) at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1/50,000 dilution was used for 1 hour at room temperature.
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Western blot analysis of Staufen on K562 cell lysates with Rabbit anti-Staufen antibody (ET1706-41) at 1/500 dilution.
Lysates/proteins at 10 µg/Lane.
Predicted band size: 63 kDa
Observed band size: 53 kDa
Exposure time: 2 minutes;
10% 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 (ET1706-41) at 1/500 dilution was used in 5% NFDM/TBST at room temperature for 2 hours. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1:200,000 dilution was used for 1 hour at room temperature.
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Western blot analysis of Staufen on LOVO cell lysates with Rabbit anti-Staufen antibody (ET1706-41) at 1/500 dilution.
Lysates/proteins at 10 µg/Lane.
Predicted band size: 63 kDa
Observed band size: 53 kDa
Exposure time: 2 minutes;
10% 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 (ET1706-41) at 1/500 dilution was used in 5% NFDM/TBST at room temperature for 2 hours. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1:200,000 dilution was used for 1 hour at room temperature.
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Immunocytochemistry analysis of HepG2 cells labeling Staufen with Rabbit anti-Staufen antibody (ET1706-41) at 1/50 dilution.
Cells were fixed in 4% paraformaldehyde for 10 minutes at 37 ℃, permeabilized with 0.05% Triton X-100 in PBS for 20 minutes, and then blocked with 2% negative goat serum for 30 minutes at room temperature. Cells were then incubated with Rabbit anti-Staufen antibody (ET1706-41) at 1/50 dilution in 2% negative goat serum overnight at 4 ℃. Alexa Fluor®488 conjugate-Goat anti-Rabbit IgG was used as the secondary antibody at 1/1,000 dilution. Nuclear DNA was labelled in blue with DAPI.
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Immunocytochemistry analysis of LOVO cells labeling Staufen with Rabbit anti-Staufen antibody (ET1706-41) at 1/50 dilution.
Cells were fixed in 4% paraformaldehyde for 10 minutes at 37 ℃, permeabilized with 0.05% Triton X-100 in PBS for 20 minutes, and then blocked with 2% negative goat serum for 30 minutes at room temperature. Cells were then incubated with Rabbit anti-Staufen antibody (ET1706-41) at 1/50 dilution in 2% negative goat serum overnight at 4 ℃. Alexa Fluor®488 conjugate-Goat anti-Rabbit IgG was used as the secondary antibody at 1/1,000 dilution. Nuclear DNA was labelled in blue with DAPI.
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Immunocytochemistry analysis of SH-SY5Y cells labeling Staufen with Rabbit anti-Staufen antibody (ET1706-41) at 1/50 dilution.
Cells were fixed in 4% paraformaldehyde for 10 minutes at 37 ℃, permeabilized with 0.05% Triton X-100 in PBS for 20 minutes, and then blocked with 2% negative goat serum for 30 minutes at room temperature. Cells were then incubated with Rabbit anti-Staufen antibody (ET1706-41) at 1/50 dilution in 2% negative goat serum overnight at 4 ℃. Alexa Fluor®488 conjugate-Goat anti-Rabbit IgG was used as the secondary antibody at 1/1,000 dilution. Nuclear DNA was labelled in blue with DAPI.
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Flow cytometric analysis of Staufen was done on SH-SY5Y cells. The cells were fixed, permeabilized and stained with the primary antibody (ET1706-41, 1/50) (red). After incubation of the primary antibody at room temperature for an hour, the cells were stained with a Alexa Fluor®488 conjugate-Goat anti-Rabbit IgG Secondary antibody at 1/1000 dilution for 30 minutes.Unlabelled sample was used as a control (cells without incubation with primary antibody; black).
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