Methods
Venous blood (20-50 mL) was obtained from patients newly diagnosed with AML prior to any treatment, from AML patients who achieved CR following initial therapy and from age-matched NC. Exosome fractions were isolated from plasma using mini-size exclusion chromatography (mini-SEC) with Sepharose 2A. Exosome protein levels, numbers and size of exosomes (qNano) and their morphology (TEM) were determined. Exosomes were characterized by Western blots for expression of exosome markers, Tsg101 and CD81, and myeloid cell-surface markers associated with AML, interleukin-3 receptor alpha chain (CD123) and C-type lectin-like molecule-1 (CLL-1), CD44 and CD96. Isolated normal human NK cells were co-incubated with AML exosomes and multiparameter flow cytometry was used to monitor changes in expression levels of NKG2D on NK cells.