Figure 5

Proliferation potential of primary keratinocytes in long-term culture and immunostaining of transplanted skin grafts. (A) ABCG2-positive keratinocytes were sorted; holoclones, meroclones and paraclones were identified, and subcultured serially on a lethally irradiated 3T3 feeder layer until their growth potential was exhausted. Triplicate cultures in each keratinocyte clones were cultured, and a total of five keratinocyte clones were analyzed. The number of cumulative population doublings was calculated using the following formula: population doublings = (log N / N ₀) / log₂, where N represents the total number of cells obtained at each passage and N ₀ represents the number of plating cells at the beginning. H, holoclone; M, meroclone; P, paraclone. The image is a typical graph. (B), (C) Immunostaining of anti-involucrin monoclonal antibody was used to detect transplanted human grafts in mouse skin samples. Positive staining (red) was observed in human grafts (single orange arrow, C), while negative staining was observed in mouse skin (double orange arrow, C), and distinct separation was observed (black arrow, B). Scale bars: B, C, 50 μm.