Reisner Y, Dagan S. of human/BALB/c chimeric mice during the first 3 weeks after PBMC transfer. Moreover, specific memory responses were elicited by vaccination with tetanus toxoid (TT) or hepatitis B virus (HBV) surface (HBs) antigen of chimeric mice transplanted with PBMC derived from TT- or HBV-immune donors. Substantially higher TT-specific B-cell frequencies were found during the first 3 weeks after vaccination in mice challenged with the specific antigen compared to the levels found in control animals. High numbers of TT-specific IFN–secreting T cells persisted in the peritoneum of vaccinated, but not of unvaccinated, animals during the entire observation period, but only low numbers of specific IL-4-secreting T cells were found in vaccinated mice. Similar results were achieved following vaccination with HBs antigen of chimeric mice, transplanted with PBMC RGFP966 of HBV immunized donors. Thus, TT or HBsAg-specific antibody responses in our model correlate closely with the existence of specific IFN–secreting T helper 1/0 cells. Furthermore, these results show that adoptive transfer of human PBMC into lethally irradiated mice provides an efficient approach to generate specific B-cell fusion partners for the production of human monoclonal antibodies and specific T-cell lines for adoptive cell therapy of malignant or infectious diseases. INTRODUCTION Severe combined immunodeficiency (SCID) mice do not reject transplants of xenogeneic cells or tissues due to a congenital lack of mature B and T cells.1 The transplantation of human peripheral blood mononuclear cells (PBMC) into SCID mice (hu-PBL-SCID mice) results in high serum levels of human immunoglobulins.2 Furthermore, specific antibody responses were generated in such chimeric mice against several bacterial, protozoal or synthetic antigens and the induction of cytotoxic T lymphocyte (CTL) RGFP966 responses has been reported in human/SCID chimeras.2C9 However, the detection of antigen-specific T-cell responses in SCID mice still represents a major problem, most likely caused by the limited engraftment of transferred human PBMC,10 progressive restriction of B- and T-cell receptor repertoires,11,12 T-cell anergy in long-term chimeras13 and the lack of professional T-cell stimulation.14 Furthermore, the frequencies of antigen-specific T cells are far too low to be detected by the conventional 3H-thymidine uptake assay.13,15,16 An alternative approach to generate human/mouse chimeras in lethally irradiated mice or rats radioprotected with SCID bone marrow (BM), was described by Lubin with TT or HBs antigen. The frequencies and cytokine patterns of the antigen-specific T helper (Th) cells and B cells induced in response to recall antigens were studied using the highly sensitive Elispot technique.23,24 Thus, we were able for the first time to quantify antigen-specific Th-cell and B-cell responses in a human/mouse chimeric model on a single cell level. Our analysis reveals a close correlation between the stimulation of strong antigen-specific Th1/0 cells and the development of high anti-TT and anti-HBs antibody levels in the serum of chimeric mice. MATERIALS AND METHODS MiceBALB/c mice (6C12-weeks-old from Olac Farms, Bicester, UK) were used as recipients of SCID BM and human PBMC. These mice were lethally irradiated by a modified split irradiation protocol as published recently (day ?4: 35 Gy; day ?1: 95 Gy).17 Bone marrow was obtained from 4C8-week-old non-obese diabetic (NOD)/SCID mice (obtained from Animal Breeding Center of the Weizmann Institute, RGFP966 Rehovot, Israel) and transplanted into recipient mice by i.v. injection of 3106 cells in 02 ml phophate-buffered saline (PBS) per mouse (day 0). NOD/SCID mice were used as they exhibit not only a lack of functional B and T cells but also a reduced natural killer (NK) cell and macrophage acitvity.25 All mice were kept under pathogen-free conditions, fed sterile food and acid water containing ciprofloxacin (20 g/ml). Preparation and transplantation of human PBMC, vaccinationPBMC were obtained from two HBV immunized donors more than 1 year after spontaneous clearance of HBV infection (serologically positive for anti-HBc and anti-HBs antibodies, negative for HBs antigen) and from two volunteers vaccinated with tetanus vaccine (RAFA, Jerusalem, Israel) several years before. All donors were healthy and tested negative for anti-hepatitis C virus (HCV) and anti-human immunodeficiency virus (HIV)1/2 antibodies. Human PBMC were isolated by leukapheresis followed by Ficoll separation. Within 1C3 days after bone marrow transplantation (BMT), 5C10107 PBMC per mouse Rabbit Polyclonal to Keratin 19 were infused intraperitoneally and mice were vaccinated with 5 g/mouse of recombinant major HBs antigen (Engerix, SKB).