TARDBP Antibody - ChIP-seq Grade

Figure 1. ChIP results obtained with the Diagenode antibody directed against TARDBP
ChIP assays were performed using K562 cells, the Diagenode antibody against TARDBP (Cat. No. C15410266) and optimized primer sets for qPCR. ChIP was performed with the “iDeal ChIP-seq” kit (Cat. No. C01010055), using sheared chromatin from 4 million cells. A titration of the antibody consisting of 1, 2 and 5 μg per ChIP experiment was analysed. IgG (1 μg/IP) was used as negative IP control. QPCR was performed with primers for the GAPDH and RFS26 promoters, used as positive controls, and for the MYOD1 gene and the Sat2 satellite repeat, used as negative controls. Figure 1 shows the recovery, expressed as a % of input (the relative amount of immunoprecipitated DNA compared to input DNA after qPCR analysis).

Figure 2. ChIP-seq results obtained with the Diagenode antibody directed against TARDBP
ChIP was performed on sheared chromatin from 5 million K562 cells using 2 μg of the Diagenode antibody against TARDBP (Cat. No. C15410266). The IP’d DNA was subsequently analysed on an Illumina HiSeq. Library preparation, cluster generation and sequencing were performed according to the manufacturer’s instructions. The 50 bp tags were aligned to the human genome using the BWA algorithm. Figure 2 shows the enrichment along the complete sequence and a 1.5 Mb region of chromosomes 3 (fig 2A and B) and 12 (fig 2C and D), respectively.

Figure 3. Western blot analysis using the Diagenode antibody directed against TARDBP
Whole cell extracts (30 μg) from Neuro2A (lane1), GL261 (lane 2), NIH3T3 (lane 3), BCL1 (lane 4) and Raw264.7 (lane 5) cells were analysed by Western blot using the Diagenode antibody against TARDBP (Cat. No. C15410266) diluted 1:3,000. The position of the protein of interest is indicated on the right; the marker (in kDa) is shown on the left.

Figure 4. Western blot analysis using the Diagenode antibody directed against TARDBP
Whole cell extracts (30 μg) from A431 (lane1), H1299 (lane 2) and HeLa (lane 3),cells were analysed by Western blot using the Diagenode antibody against TARDBP (Cat. No. C15410266) diluted 1:1,000. The position of the protein of interest is indicated on the right; the marker (in kDa) is shown on the left.

Figure 5. Western blot analysis using the Diagenode antibody directed against TARDBP
Whole cell extracts (30 μg) from HeLa cells transfected with sh-TARDBP (lane1), or a mock control (lane 2) were analysed by Western blot using the Diagenode antibody against TARDBP (Cat. No. C15410266) diluted 1:1,000. The position of the protein of interest is indicated on the right; the marker (in kDa) is shown on the left. The lower panel shows the signal obtained with an ACTB antibody, used as a loading control.

Figure 6. Immunoprecipitation using the Diagenode antibody directed against TARDBP
Immunoprecipitation was performed on whole cell extracts from HeLa cells using 2 μg of the Diagenode antibody against TARDBP (Cat. No. C15410266). An equal amount of rabbit IgG was used as a negative control. The immunoprecipitated TARDBP protein was detected by western blot with the TARDBP antibody diluted 1:1,000. The IP with the TARDBP antibody and with the IgG negative control are shown in lane 3 and lane 2, respectively. Lane 1 shows the input (40 μg of HeLa whole cell extract).

Figure 7. Immunofluorescence with the Diagenode antibody directed against TARDBP
HeLa cells were fixed with 4% formaldehyde for 15’ at room temperature and stained with the Diagenode antibody against TARDBP (Cat. C15410266) diluted 1:200 (left). The right picture shows costaining with Hoechst 33342 nucleic acid stain.

Figure 8. Immunohistochemistry using the Diagenode antibody directed against TARDBP
Formalin fixed paraffin embedded rat brain tissue (figure A) or Cal27 Xenograft (figure B) was stained with the Diagenode antibody against TARDBP (Cat. No. C15410266) diluted 1:500 and 1:100, respectively, followed by a peroxidase labelled goat anti-rabbit secondary antibody.