Fig 1: TDP-43 antibody screening by immunofluorescence.HAP1 WT and TARDBP KO cells were labelled with a green or a far-red fluorescent dye, respectively. WT and KO cells were mixed and plated to a 1:1 ratio in a 96-well plate with a glass bottom. Cells were stained with the indicated TDP-43 antibodies and with the corresponding Alexa-fluor 555 coupled secondary antibody including DAPI. Acquisition of the blue (nucleus-DAPI), green (identification of WT cells), red (antibody staining) and far-red (identification of KO cells) channels was performed. Representative images of the merged blue and red (grayscale) channels are shown. WT and KO cells are outlined on both channels with green and magenta dashed lines, respectively. Antibody dilutions were chosen according to the recommendations of the antibody supplier. Exceptions were given for antibodies 12892-1-AP, MA5-32627**, A19123**, 89789**, 89718** and ab133547** which were titrated to 1/400, 1/1000, 1/900, 1/30, 1/10 and 1/700, respectively, as the signals were too weak when following the supplier’s recommendations When the concentration was not indicated by the supplier, which was the case for antibodies 80001-1-RR**, GTX630196* and ab254166**, we tested antibodies at 1/700, 1/1000 and 1/500, respectively. At these concentrations, the signal from each antibody was in the range of detection of the microscope used. Antibody dilution used: 10782-2-AP at 1/400, 12892-1-AP at 1/250, 800001-1-RR** at 1/700, 80002-1-RR** at 1/250, MAB7778* at 1/500, NBP1-92695* at 1/1000, 711051** at 1/500, MA5-27828* at 1/1000, MA5-32627** at 1/1000, A19123** at 1/900, 89789** at 1/30, 89718** at 1/10, GTX630196* at 1/1000, GTX630197* at 1/1000, ab109535** at 1/30, ab133547** at 1/700, ab190963** at 1/800, ab254166** at 1/500. Bars = 10 μm. *Monoclonal antibody, **Recombinant antibody.
Fig 2: TDP-43 antibody screening by Western blot.Lysates of HAP1 (WT and TARDBP KO) were prepared and 50 μg of protein were processed for Western blot with the indicated TDP-43 antibodies. The Ponceau stained transfers of each blot are shown. Antibody dilutions were chosen according to the recommendations of the antibody supplier. Exceptions were given for antibody 80001-1-RR**, which was titrated to 1/1000 as the signal was too weak when following the supplier’s recommendations. When the concentration was not indicated by the supplier, which was the case for antibody 80002-1-RR**, we tested the antibody at 1/1000. Antibody dilution used: 10782-2-AP at 1/5000, 12892-1-AP at 1/1000, 800001-1-RR** at 1/1000, 80002-1-RR** at 1/1000, MAB7778* at 1/500, NBP1-92695* at 1/1000, 711051** at 1/1000, MA5-27828* at 1/1000, MA5-32627** at 1/1000, A19123** at 1/1000, 89789** at 1/1000, 89718** at 1/1000, GTX630196* at 1/500, GTX630197* at 1/500, ab109535** at 1/2000, ab133547** at 1/1000, ab190963** at 1/1000, ab254166** at 1/1000. Predicted band size: 45 kDa. *Monoclonal antibody, **Recombinant antibody.
Fig 3: TDP-43 antibody screening by immunoprecipitation.HAP1 lysates were prepared, and IP was performed using 2.0 μg of the indicated TDP-43 antibodies pre-coupled to Dynabeads protein G or protein A. Samples were washed and processed for Western blot with the indicated TDP-43 antibody. For Western blot, 80002-1-RR** was used at 1/1000. The Ponceau stained transfers of each blot are shown for similar reasons as in Figure 1. SM=4% starting material; UB=4% unbound fraction; IP=immunoprecipitate. *Monoclonal antibody, **Recombinant antibody.
Supplier Page from Abcam for Anti-TDP43 antibody [DB9]