PMC5796899_01
Mean fluorescence intensity of anti-IgG coupled to Phycoerythrin staining after incubation of patient serum with the indicated drug on monocytes, lymphocytes, and neutrophils from patient (A) or a healthy control (B). (C) Gating strategy allowing the identification of natural killer (NK) cells (CD3-/4-/14-/19-/56+ cells). (D) Concatenation of FACS plot showing NK cells activation markers after NK cells culture in vitro in the indicated conditions. Results are expressed in percentage of positive cells for each condition.
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F1
We observed that the incubation of patient serum with rasburicase leads to a specific IgG staining on monocytes and lymphocytes but not on neutrophils, while IgG MFI was not increased in the presence of allopurinol (Figure 1A). This phenomenon was also observed, albeit at a lower extent, when patient serum was incubated with rasburicase and leukocytes from a healthy donor (Figure 1B). Navios instrument (Beckman Coulter, Brea, CA, USA), and data were analyzed with FlowJo (Treestar, Ashland, OR, USA). NK cells were defined as CD3-/4-/19-CD56+ cells as shown in Figure 1C. Under these conditions, NK cells degranulated (increase of the CD107a expression) and produced high amount of MIP-1beta similar to positive control with K562 cells (Figure 1D), whereas the production of IFN-gamma was low in all conditions. By contrast, when PBMCs were incubated with rasburicase without patient serum [but with 10% fetal calf serum (FCS)] or with patient serum but without any drug or with allopurinol, no sign of NK cells activation was observed (Figure 1D).