The Journal of Experimental Medicine
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© The Rockefeller University Press, 0022-1007/1996/12/2217/ $5.00
The Journal of Experimental Medicine, Volume 184, Number 6, December 1, 1996 2217-2230

Articles

Ordering of Human Bone Marrow B Lymphocyte Precursors by Single-Cell Polymerase Chain Reaction Analyses of the Rearrangement Status of the Immunoglobulin H and L Chain Gene Loci

Paolo Ghia*, Edwin ten Boekel*, Eva Sanz{ddagger},§, Antonio de la Hera{ddagger},§, Antonius Rolink*, and Fritz Melchers*

From the * Basel Institute for Immunology, CH-4005 Basel, Switzerland; {ddagger} Centro de Investigaciones Biologicas, E-28006 Madrid, Spain; and § Department of Medicine, Alcala University, E-28871 Madrid, Spain

CD19+CD10+ human B lineage bone marrow cells were separated into cycling or resting cells, which differ in their expression of CD34, VpreB, recombination activating gene (RAG-1), and terminal deoxynucleotidyl transferase (TdT). Polymerase chain reaction analyses developed for DHJH and V{kappa}J{kappa}, V{kappa}J{kappa}K(de) and V{kappa}K(de) rearrangements with DNA of single cells and a comparison with B lineage cell development in mouse bone marrow, allow to delineate the human B lymphocyte pathway of development as follows: CD34+VpreB+RAG-1+TdT+, DHJH-rearranged, {kappa}L germline cycling pre-B I cells -> CD34–VpreB+µH chain+ (pre-B receptor+) RAG-1–TdT–, VHDHJH-rearranged, {kappa}L germline, cycling pre-B II cells -> CD34–VpreB–, intracytoplasmic µH chain+ (pre-B receptor–) RAG-1+/– TdT–, VHDHJH-rearranged, mainly {kappa}L germline cycling pre-B II cells -> CD34–VpreB– intracytoplasmic µH chain+, RAG-1+TdT–, VHDHJH-rearranged, V{kappa}J{kappa}-rearranged, IgM–, resting pre-B II cells CD34+VpreB–, sIgM+, RAG-1+TdT–, VHDHJH- and V{kappa}J{kappa}-rearranged IgM+ immature B cells -> CD34–, CD10–, sIgM+/sIgD+ mature B cells. This order, for the first time established for human B lineage cells, shows striking similarities with that established for mouse B lineage cells in bone marrow.

Address correspondence to Dr. Fritz Melchers, Basel Institute for Immunology, Grenzacherstrasse 487, CH4005 Basel, Switzerland.

The Basel Institute for Immunology was founded and is supported by F. Hoffmann-La Roche Ltd., Basel, Switzerland. E. Sanz was supported by contracts from the CSIC and grant CAM92/126, and A. de la Hera was supported by grants SAF-93-0925 and SAF-96-0201 from the CICY.

1Abbreviations used in this paper: BIO, biotin; H, heavy; {kappa}de, {kappa}-deleting element; L, light; RAG, recombination activating gene; RT, reverse transcription; s, surface; Tdt, terminal deoxynucleotidy transferase.


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