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Western blot analysis of LFNG on different lysates with Rabbit anti-LFNG antibody (HA721367) at 1/1,000 dilution.
Lane 1: Hela cell lysate (15 µg/Lane)
Lane 2: K-562 cell lysate (15 µg/Lane)
Lane 3: HL-60 cell lysate (15 µg/Lane)
Lane 4: MCF7 cell lysate (15 µg/Lane)
Lane 5: HT-29 cell lysate (15 µg/Lane)
Lane 6: C2C12 cell lysate (15 µg/Lane)
Lane 7: PC-12 cell lysate (15 µg/Lane)
Lane 8: Mouse testis tissue lysate (30 µg/Lane)
Predicted band size: 42 kDa
Observed band size: 42 kDa
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 (HA721367) at 1/1,000 dilution was used in 5% NFDM/TBST at room temperature for 2 hours. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1:100,000 dilution was used for 1 hour at room temperature.
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Immunocytochemistry analysis of HUVEC cells labeling LFNG with Rabbit anti-LFNG antibody (HA721367) at 1/500 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-LFNG antibody (HA721367) at 1/200 dilution in 2% negative goat serum overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor™ 488, HA1121) was used as the secondary antibody at 1/1,000 dilution. Nuclear DNA was labelled in blue with DAPI.
Beta tubulin (M1305-2, red) was stained at 1/200 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (iFluor™ 594, HA1126) was used as the secondary antibody at 1/1,000 dilution.
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Immunocytochemistry analysis of MCF7 cells labeling LFNG with Rabbit anti-LFNG antibody (HA721367) at 1/500 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-LFNG antibody (HA721367) at 1/200 dilution in 2% negative goat serum overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor™ 488, HA1121) was used as the secondary antibody at 1/1,000 dilution. Nuclear DNA was labelled in blue with DAPI.
Beta tubulin (M1305-2, red) was stained at 1/200 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (iFluor™ 594, HA1126) was used as the secondary antibody at 1/1,000 dilution.
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Flow cytometric analysis of K-562 cells labeling LFNG.
Cells were fixed and permeabilized. Then stained with the primary antibody (HA721367, 1ug/ml) (red) compared with Rabbit IgG Isotype Control (green). After incubation of the primary antibody at +4℃ for an hour, the cells were stained with a iFluor™ 488 conjugate-Goat anti-Rabbit IgG Secondary antibody (HA1121) 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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