Dose response and time course analysis of MK signaling after TPO stimulation. Dose response (A) and time-course (B) analysis of MK signaling in response to TPO. Mature MKs derived from BM cultures of control and JAK2V617F animals were purified by means of a BSA gradient and starved of cytokines for 4 hours. They were then stimulated with the dose of TPO indicated and for various lengths of time. Cell lysates were subsequently analyzed by western blotting for evidence of signaling downstream of JAK2. Densitometry analysis confirmed a significant increase in phosphorylation of both ERK and STAT3 in response to TPO in JAK2V617F MKs. Error bars represent SD.