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. 2016 Apr 25;11(4):e0152073. doi: 10.1371/journal.pone.0152073

Fig 4. Detection of CD200 in membrane extracts from PMA-treated CLL.

Fig 4

Membrane proteins from aliquots of treated and untreated cells from experiments shown in Fig 3 were extracted and analyzed by a) ELISA, and b) Western blotting for CD200. Both methods showed loss of CD200 from the membrane fraction in PMA-treated cells from patients that responded to PMA stimulation as determined by FACS (patients not shedding CD200 in response to PMA as assessed by FACS are highlighted in italic). c) CLL cells from another cohort of patients (n = 3) were cultured in AIMV medium, with or without PMA stimulation, and treated with 50μg/ml of TAPI-0. Cells were harvested at 24-hr and stained for CD200 and CD19. TAPI-0 restored cell surface expression of CD200 on PMA-treated cells from patients 139 and 158 (2 upper panels). CLL cells from patient 16 showed no response (attenuation of PMA-induced sCD200 shedding) to TAPI-0 with PMA causing only a low level of CD200 shedding in this case (bottom panel). d) %PMA-induced shedding, and %inhibition of PMA-induced shedding by TAPI-0 for the three patients shown in panel d).