IntroductionThe risk of transfusion-transmitted hepatitis B virus (HBV) infection has been reduced by screening all blood donations for HBV surface antigen (HBsAg) since 1970. It was generally accepted that the disappearance of HBsAg indicates the clearance of HBV. Meanwhile, many reports on positive findings for HBV DNA in the liver and blood of HBsAg-negative individuals positive for antibodies against HBV core antigen (anti-HBc) and/or HBsAg (anti-HBs) have been published. 1-8 Blum et al described, in a patient with HBsAg-negative chronic hepatitis who was positive for anti-HBc, anti-HBs, and antibodies against HBe antigen (anti-HBe), a latent HBV infection in hepatocytes with extrachromosomal presence of a full-length viral genome. 9 Michalak et al demonstrated the long-term persistence of HBV DNA in serum and peripheral blood mononuclear cells of patients up to 70 months after complete clinical, biochemical, and serologic recovery from acute viral hepatitis. 10 Rehermann et al showed that traces of HBV were often detectable in the blood for many years after clinical recovery from acute hepatitis, despite the presence of serum antibodies and HBV-specific cytotoxic T lymphocytes (CTLs). 11 These findings suggest that sterilizing immunity to HBV frequently fails to occur and that traces of virus can maintain the CTL response for decades, apparently creating a negative feedback loop that keeps the virus under control, perhaps for life. This was supported by Pasquetto et al, who showed that cytoplasmic HBV nucleocapsids and their cargo of replicative DNA intermediates survive CTL-induced apoptosis of hepatocytes in vitro, 12 and by other groups that demonstrated ongoing viral replication in the liver tissue of patients and healthy individuals after loss of HBsAg. [13][14][15] Furthermore, reactivation of apparently cured HBV infection has been described under chemotherapy or immunomodulating therapy after renal and bone marrow transplantation, and in some of these cases a reverse seroconversion from anti-HBs to HBsAg has been observed. [16][17][18][19] The residual risk of posttransfusion HBV infection has been calculated by several groups in the United States and Germany on the basis of HBV incidence data and the duration of the early window period until HBsAg becomes detectable to be 1:63 000 and less than 1:100 000 blood donations, respectively. 20,21 It has been shown that blood donations of HBsAg-and anti-HBs-negative but anti-HBc-positive HBV carriers can cause posttransfusion hepatitis B. 22;23 Thus, Mosley et al suggested that anti-HBc screening of blood donations might prevent HBV transmission from HBsAgnegative blood donors and that donors positive for anti-HBs as well should be considered noninfectious for HBV. 24 The feasibility of routine polymerase chain reaction (PCR) screening of blood donations in a blood bank setting has been shown by Roth et al. 25 In Germany, the Paul Ehrlich Institute (PEI, Langen, Germany), the institute that defines German Drug Law and is responsible for specific regulati...

The actual frequency of antibody formation in our patients is much lower than assumed. On the other hand, prolonged hemolysis probably induced by additional autoreactive antibodies might occur. This possible complication has not yet been addressed. Further studies might reveal whether a less restricted transfusion policy with respect to D matching is justified in selected patients.

Summary. Natural killer (NK) cells are assumed to contribute to a graft-versus-leukaemia effect. In vitro experiments have shown that many leukaemic cells are NK-cell sensitive. Nevertheless, no data concerning the influence of purified NK cells on malignant myeloma (MM) cells exist. We co-incubated NK cells with three different MM cell lines and fresh bone marrow samples of nine MM patients. The proportion of vital MM cells was determined before and after co-cultivation by a flow-cytometry-based assay. All MM cells tested, with the exception of one cell line (NCI H929), were susceptible to a NK-cell attack even without exogenous interleukin 2 (IL-2). The mean killing of the native MM samples was 23AE1 ± 5AE4% and 34AE5 ± 6AE5% at 10:1 and 20:1 effector:target ratio respectively, This corresponded to about 2/3 of those values obtained with the highly sensitive line K562. In contrast, CD34-positive haematopoietic stem cells as well as peripheral mononuclear cells were completely resistant under similar experimental conditions (1AE3% killing). To elucidate the underlying triggering mechanisms, we measured human leucocyte antigen (HLA)-class I expression of the MM cells. No evidence for HLA loss, which could have explained the NK-cell recognition if it occurred, was demonstrated. These findings may contribute to the understanding of in vivo NK-cell activation and encourage clinical applications of NK cells for MM patients.

Natural killer (NK) lymphocytes can be used for adoptive immunotherapeutic strategies. Alternatively, they may be employed as adjuvants for stem cell/bone marrow transplantation, either to re-induce remission, or to purge autografts of contaminating malignant cells. We developed a new protocol that enables the generation of NK cells on a clinical scale in a closed system that enables good manufacturing practice (GMP) conformity. Aside from the initial NK cell inoculum, our protocol includes activated feeder cells [irradiated peripheral blood mononuclear cells (PBMC) and no transformed blasts], cytokines [interleukin-2 (IL-2) and IL-15], human serum, and a complex basic media formulation. During the whole expansion period of approximately 14 days, the cells were handled in PTFE (Teflon) bags, whereby fresh medium was added without opening the system. The use of immortalized or virus-transformed feeder cells, as used in many other current research protocols, was completely avoided. A precise controlling of a number of environmental factors was necessary to achieve reproducible results. Increases in NK cell number ranged between 80- and 200-fold. The resulting NK cells were CD56(+), CD3(-), and CD16(+) (75%). They were highly cytotoxic against different malignant target cells and did not produce significant levels of interferon-gamma. Therefore, they belonged to the cytotoxic rather than the immunoregulatory NK subpopulation. No non-specific activation against normal allogenous lymphocytes occurred. This work might permit the realization of future protocols for evaluating the clinical effect of NK lymphocytes in human disease.