Figure 2 | Scientific Reports

Figure 2

From: Development of a novel immunoassay to detect interactions with the transactivation domain of p53: application to screening of new drugs

Figure 2

Development of AlphaLISA assay to detect recombinant protein p53-His in cellular extracts. (a) A matrix cross-titration of a panel of cell-extract amounts against a dilution series of biotinylated anti-His antibodies revealed the optimal cell-extract amount for subsequent experiments. (b) The incubation time of acceptor beads, the biotinylated anti-His antibodies and cell extracts was tested between 5 and 25 min. AlphaLISA signal (S/B) is represented as the ratio of AlphaLISA signal in the presence and absence of cell extracts. For subsequent experiments an incubation time of 20 min (S/B = 202) was adopted. (c) Titration of biotinylated anti-His antibody with the optimal amount of cell extracts and incubation time. (d,e) Titration of AlphaLISA acceptor and donor beads at previously determined optimal conditions. (f) Dose-response curve of cell extracts containing p53-His. Each point is an average of three independent experiments.

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