NAT10 Recombinant Rabbit Monoclonal Antibody [JE55-38]

Western blot analysis of NAT10 on different lysates with Rabbit anti-NAT10 antibody (ET7111-23) at 1/2,000 dilution.

Lane 1: HL-60 cell lysate
Lane 2: Daudi cell lysate
Lane 3: Mouse stomach tissue lysate
Lane 4: Rat colon tissue lysate
Lane 5: Rat brain tissue lysate

Lysates/proteins at 20 µg/Lane.

Predicted band size: 116 kDa
Observed band size: 116 kDa

Exposure time: Lane 1-2: 8 seconds; Lane 3-5: 3 minutes; ECL: K1802;
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 (ET7111-23) at 1/2,000 dilution was used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1/50,000 dilution was used for 1 hour at room temperature.

☑ Knockdown (KD)

Western blot analysis of NAT10 on different lysates with Rabbit anti-NAT10 antibody (ET7111-23) at 1/500 dilution.

Lane 1: Hela-si NT cell lysate (10 µg/Lane)
Lane 2: Hela-si NAT10 cell lysate (10 µg/Lane)

Predicted band size: 116 kDa
Observed band size: 116 kDa

Exposure time: 1 minute;
4-20% SDS-PAGE gel.

ET7111-23 was shown to specifically react with NAT10 in Hela-si NT cells. Weakened band was observed when Hela-si NAT10 sample was tested. Hela-si NT and Hela-si NAT10 samples were subjected to SDS-PAGE. Proteins were transferred to a PVDF membrane and blocked with 5% NFDM in TBST for 1 hour at room temperature. The primary antibody (ET7111-23, 1/500) and Loading control antibody (Rabbit anti-GAPDH, ET1601-4, 1/10,000) were used in 5% BSA 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.

Immunocytochemistry analysis of Daudi cells labeling NAT10 with Rabbit anti-NAT10 antibody (ET7111-23) at 1/250 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-NAT10 antibody (ET7111-23) at 1/250 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor™ 488, HA1121) 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 (HA601187, red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (iFluor™ 594, HA1126) was used as the secondary antibody at 1/1,000 dilution.

Immunohistochemical analysis of paraffin-embedded human lung carcinoma tissue using anti-NAT10 antibody. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) (high pressure) for 2 minutes. The tissues were blocked in 5% BSA for 30 minutes at room temperature, washed with ddH₂O and PBS, and then probed with the primary antibody (ET7111-23, 1/50) for 30 minutes 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.

Immunohistochemical analysis of paraffin-embedded human colon carcinoma tissue using anti-NAT10 antibody. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) (high pressure) for 2 minutes. The tissues were blocked in 5% BSA for 30 minutes at room temperature, washed with ddH₂O and PBS, and then probed with the primary antibody (ET7111-23, 1/50) for 30 minutes 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.

Immunohistochemical analysis of paraffin-embedded mouse brain tissue using anti-NAT10 antibody. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) (high pressure) for 2 minutes. The tissues were blocked in 5% BSA for 30 minutes at room temperature, washed with ddH₂O and PBS, and then probed with the primary antibody (ET7111-23, 1/200) for 30 minutes 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.

Immunohistochemical analysis of paraffin-embedded rat brain tissue with Rabbit anti-NAT10 antibody (ET7111-23) 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 (ET7111-23) 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.