Figure 2.

General experimental setup of the original HIMA [10]. Primary HUFs are isolated and grown under standard culture conditions (20% oxygen). For the HIMA primary HUFs are seeded on six-well plates (P1), treated with a mutagen and serially passaged under standard culture conditions (20% oxygen) until cells undergo senescence (P5+). Mutated cells eventually emerge from senescence and can be expanded into immortalised clonal cell lines (P8+). TP53 sequencing with isolated DNA from all clones is performed to identify TP53 mutations and to evaluate the pattern of mutations induced by the mutagen.