PIA assay for FLT3. This assay serves as a validated surrogate for in vivo FLT3 inhibition in patients treated with FLT3 inhibitors. Whole blood is collected at a trough time point from patients treated with an FLT3 inhibitor, preferably when the drug is at steady-state. An FLT3-ITD–expressing cell line, such as Molm14, is then incubated in the plasma, and the phosphorylation status of FLT3 is analyzed by immunoblotting. The results are normalized to control or pretreatment plasma. The immunoblots can be quantitated by densitometry. The blots are from a patient treated with 80 mg/d or 60 mg/d of lestaurtinib, a first-generation FLT3 inhibitor.¹⁰³  Professional illustration by Somersault18:24.