CXCL12 treatment results in prolonged activation of ERK in ZAP-70⁺ but not ZAP-70^– CLL B cells. (A) Lysates were collected from CLL B cells treated for varying times with CXCL12 (100 ng/mL). Western blots were probed with an anti–phospho-p44/42 ERK (P-ERK) antibody (top panel) and then with an anti-p44/42 ERK (T-ERK) antibody to demonstrate loading (bottom panel). (i) ZAP-70⁺ PB CLL cells. (ii) ZAP-70^– PB CLL cells. (B) Densitometric quantification of P-ERK and T-ERK Western blots from 7 ZAP-70⁺ and 7 ZAP-70^– patients was carried out and the ratio of P-ERK to T-ERK was calculated to normalize for loading. Results are presented as mean ± SEM (Student 2-sample 2-tailed t test). (C) ERK activation following addition of CXCL12 in ZAP-70⁺ CLL B cells is sustained for up to 2 hours.