Eur. J. Immunol. 1988.18: 1299-1302 CR2 receptors on thymocytes 1299 zy Short paper Constantine D. TsoukasoA and John D. Lambris' Department of Biology and Molecular Biology Institute', San Diego State UniversitySan Diego, Department of Basic and Clinical Research', Research Institute of Scripps Clinic, La Jolla and Basel Institute for Immunology', Base1 1 Introduction Expression of CR21EBV receptors on human thymocytes detected by monoclonal antibodies* The biologic effects of the third component of complement, C3, are mediated via receptors which specifically bind the enzymatic degradation products resulting from the cleavage of C3. One of the products, C3d, has been associated with binding to the second complement receptor CR2 (CD21). This receptor, which is identical to the receptor for Epstein-Barr virus (EBV), has been primarily found on cells of the B lineage, but not on mature T cells or other cells of erythroid or myeloid lineages. In the present investigation, we report the presence of CR2 on human thymocytes. Indirect immunofluorescence analysis employing monoclonal anti-CR2 antibodies revealed a range of thymocyte reactivity from 15% to 63% in thirteen experiments using cells of different donors. Reactivity was always greater with the monoclonal anti-CR2 (CD21) antibody HB-5 than with two other antibodies which recognize distinct epitopes on the CR2 molecule. Two-color immunofluorescence analysis indi- cated that the brightest of the HB-5-stained thymocytes also reacted with the mono- clonal anti-CD1 antibody T6 (immature thymocyte marker) while some of the duller HB-5-staining cells reacted with the monoclonal anti-CD3 antibody Leu-4 (mature thymocyte marker). Immunoprecipitation of CR2 on thymocytes with antibody HB-5 and polyacrylamide gel electrophoretic analysis revealed a protein of 145 kDa molecular mass which is consistent with the size of CR2 found on B lymphocytes. These findings raise several questions regarding the biologic role of CR2-EBV recep- tor on cells of the T lineage. The third component of complement, C3, is cleaved during the activation cascade to various fragments which bind to distinct cell surface receptors [l-31. One of these fragments, C3d, has been associated with binding to the second complement recep- tor CR2 (CD21) [l-41. This receptor is present on cells of the B lineage, but absent from peripheral T cells, natural killer cells and cells of erythroid or myeloid lineages [5]. It has been reported that on B cells CR2 is identical to the receptor for Epstein-Barr virus (EBV) [6-81. zyxwvuts In the present investigation, we have used indirect immunofluorescence and immuno- precipitation techniques with anti-CR2 monoclonal antibodies (mAb) to demonstrate the presence of CR2 on human thymo- cytes. 2 Materials and methods 2.1 mAb The mAb used in this study, and their reactivities and sources are listed in Table 1. All antibodies were purified as previously [I 67501 zyxwvu * This work was supported in part by NIH grants GM39518, AR36010, AI21014, and RR00833. + The Basel Institute for Immunology was founded and is entirely supported by F. Hoffmann-La Roche, Ltd, Co., Basel, Switzerland. This is publication number 5175BCR from the Research Institute of Scripps Clinic. Correspondence: Constantine Tsoukas, Department of Biology and Molecular Biology Institute, San Diego, CA 92182, USA Abbreviations: CR2: Complement receptor type 2 EBV: Espstein- Barr virus FSC: Forward scatter mAb: Monoclonal antibody- (ies) PE: Phycoerythnn SDS-PAGE: Sodium dodecyl sulfate-poly- acrylamide gel electrophoresis Table 1. Reactivity of mAb used in this study mAb') Reactivityb) Source') OKT6 T6-RDld) OKTll 64.1 Leu-4-PEe) Leu-12 and L.cu-T~-PE" HB-5 antiB2 OW7 CD1 CD1 CD2 CD3 CD3 CD19 CD21 CD21 CD21 ATCC Coulter ATCC Genetic Systems Becton Dickinson Becton Dickinson ATCC Coulter Ortho Antibodies OKT6, Leu-4, and Leu-12 are of the IgG, isotype; OKTl1,64.1, HB-5, and OKB7 are IgG,,; anti-B2 is an IgM anti- body. The CD nomenclature is according to the 3rd International Work- shop on Human Leukocyte Differentiation Antigens [ lo]. ATCC, American Type Culture Collection (Rockville, MD); Coulter Immunology (Hialeah, FL); Genetic Systems (Seattle, WA); Becton Dickinson (Mountain View, CA); Ortho Diagnostic Systems (Raritan, NJ). T6-RD1, PE-conjugated T6. Leu-rl-PE, Leu-12-PE, PE-conjugated Leu-4 and Leu-12. described zyxwv [9]. Fluorescein-conjugated goat anti-mouse immunoglobulin (Ig) was purchased from Jackson Irnmuno- research (West Grove, PA). 2.2 Cell source Thymus glands, obtained from pediatric patients undergoing corrective cardiac surgery, were minced through a stainless steel screen and the cells were isolated by Ficoll-Hypaque (Pharmacia, Uppsala, Sweden) fractionation [ 111. 0 VCH VerlagsgesellschaftmbH, D-6940 Weinheim, 1988 0014-2980/88/0808-1299$02.50/0