Characterization of Two Novel mAbs Recognizing Different Epitopes on CD43

JL1, a specific epitope on CD43, is a potential biomarker for the diagnosis of acute leukemia. Although qualitative assays for detecting leukemia-specific CD43 exist, there is a need to develop quantitative assays for the same. Here, we developed two novel monoclonal antibodies (mAbs), 2C8 and 8E10, recognizing different epitopes on CD43. These clones are capable of pairing with YG5, another mAb against JL1 epitope, because they were selectively obtained using sandwich ELISA. Antigens recognized by 2C8 and 8E10 were confirmed as CD43 by western blotting using the CD43-hFC recombinant protein. When expression on various leukemic cell lines was investigated, 2C8 and 8E10 displayed a disparity in the distribution of the epitope. Enzyme assays revealed that these mAbs recognized a sialic acid-dependent epitope on CD43. Using normal thymus and lymph node paraffin-embedded tissues, we confirmed a difference in the epitopes recognized by the two mAbs that was predicted based on the maturity of the cells in the tissue. In summary, we developed and characterized two mAbs, 2C8 and 8E10, which can be used with YG5 in a sandwich ELISA for detecting leukemia-specific CD43.

Functional and Phenotypic Characterization of Cord Blood-Derived Dendritic Cells / 대한소아혈액종양학회지

PURPOSE: Hematopoietic stem cells from umbilical cord blood are one of the useful resources for stem cell transplantation in the various adult and childhood diseases. Immunologic complications of transplantation, e.g., graft-vs-host disease, occur much less with transplantation of cord blood stem cells. Cord blood-derived dendritic cells (CB-DCs) are known to be different from adult peripheral blood-derived dendritic cells (PB-DCs) in immunologic characteristics. These phenomena might be related to the characteristics of hematopoietic cells in cord blood. Therefore, we analysed characteristics of dendritic cells, which are well-known immune-provoking cells, derived from cord blood precursors. METHODS: Dendritic cells were differentiated from plastic-adherent cord blood monocytes in the presence of GM-CSF and IL-4. Immunophenotype was analysed by flow cytometry and expression of IDO (indoleamine 2, 3-dioxygenase), an enzyme expressed in immune-regulating or tolerogenic DCs, IL-12, IL-10 and IL-6 was measured by RT-PCR along in vitro differentiation. Changes in expression of cytokines and IDO after antibody engagement were also analysed. RESULTS: CB-DCs were very similar to PB-DCs in immunophenotype and expression of cytokines. But CB-DCs expressed IDO transcripts much earlier than PB-DCs during differentiation from precursors. Engagement of CB-DCs with DU-1 mAb induced upregulation of IDO and downregulation of IL-6. CONCLUSION: Although immunophenotype and cytokine expression pattern of CB-DCs were quite similar to those of PB-DCs, CB-DCs expressed IDO earlier than PB-DCs. This might be related to the phenomena that CB-DCs are less immunogenic or, sometimes, tolerance-inducing.