We have investigated CD52 expression and alemtuzumab (Campath-1H®) mediated lysis in acute lymphoblastic leukaemia (ALL) cell lines and freshly isolated samples. CD52 was expressed in 4 out of 9 ALL cell lines studied (44%), only two of which expressed it at high levels: the preB ALL REH carrying a t(12;21) translocation and the B-ALL Silti with rearranged c-myc (both with MFI of about 270). In contrast, of 57 freshly isolated ALL samples analysed, 86% of cases expressed CD52, albeit at varying levels. CD52 expression was generally more frequent and more intense on more mature ALL along either the B or T lineage. In particular none of the 4 proB-ALL carrying a t(4;11) translocation expressed CD52 (0%), whereas 100% of B linage ALL with a t(12;21), t(9;22) or t(1;19) translocations (total 19 cases analysed) stained positive for CD52 at varying levels (100%). We next analysed alemtuzumab and complement mediated lysis (CDC) and ADCC. ADCC was equivalent in different CD52+ lines, reaching 50% lysis at a 60:1 effector:target ratio. In contrast complement dependent cytotoxicity was variable in different cell lines, since the REH cell line bearing the t(12;21) translocation showed 47% lysis at 10 μg/ml alemtuzumab compared to 0–6% for the other cell lines (Silti and the B lymphoma EsIII), expressing equivalent amounts of CD52. Interestingly amongst freshly isolated ALL, 10/10 t(12;21) ALL samples showed very high CDC (mean 96% lysis) in presence of 10 μg/ml alemtuzumab and human serum, compared to the other 15 CD52+ cases tested in the same conditions which showed a mean of 19% lysis (p<0.0001). Furthermore in t(12;21) samples, near maximal CDC was obtained with as low as 1 μg/ml alemtuzumab whereas other ALL cases required 10 to 100 μg/ml alemtuzumab for significant lysis to be observed. CDC correlated only in part with CD52 levels, suggesting that CD52 expression as well as other yet undefined factors contribute to the particular sensitivity of t(12;21) cells. In non t(12;21) ALL cases showing low lysis, CDC could be increased 2 to 3 fold by increasing alemtuzumab concentration to 100 μg/ml, or by blocking the CD55 and CD59 complement inhibitors. Furthermore, increased cytotoxicity (up to 100%) could be obtained in several non t(12;21) ALL cases by combining alemtuzumab with fludarabine or mafosfamide, with an approximately additive effect observed between the antibody and these chemotherapeutic drugs. We conclude that the cytotoxic activity of alemtuzumab against CD52+ ALL is variable and its effectivenes can be improved by the combination with cytotoxic drugs.