Background: Hemophagocytic lymphohistiocytosis (HLH) is a serious immune disorder. Epstein-Barr virus (EBV) is the most common trigger of secondary HLH. T-cell receptor (TCR) gene rearrangement is detectable in half of patients with EBV-associated HLH using Southern blotting and conventional polymerase chain reaction (PCR) analyses. However, its clinical significance is unclear. The impact of TCR gene clonality on the outcome of patients with EBV-HLH should be evaluated using more sensitive methods. In this study, we used BIOMED-2 multiplex PCR and GeneScan analysis to evaluate TCR gene rearrangement in childhood EBV-HLH. Methods: Six children with EBV-HLH were enrolled in this study. EBV load in blood was quantified by real-time PCR. TCR gene rearrangement was analyzed by BIOMED-2 multiplex PCR. Results: All 6 patients showed monoclonal peaks in TCR beta and/or TCR gamma at diagnosis. Serial monitoring of one patient disclosed a change in the rearrangement pattern of the TCR genes in response to immunochemotherapy. Conclusion: These findings suggest that BIOMED-2 multiplex PCR, a highly sensitive method for detecting T-cell clonality, is useful for predicting the therapeutic response in childhood EBV-HLH. (C) 2011 Elsevier B.V. All rights reserved.