DIgR2 is a membrane receptor preferentially expressed by APCs. (A) Northern blots showing the expression of several DIgR2 transcripts in immune and nonimmune tissues. (B) RT-PCR analysis of various mouse cell lines (left) and freshly isolated cells (right) demonstrating restricted expression of DIgR2 in B cells and myeloid cells. (C) Quantitative RT-PCR (Q-RT-PCR) analysis of DIgR2 levels during BM-DC culture. The relative expression of DIgR2 mRNA in the cultured cells was determined daily by real-time quantitative PCR. On day 7 of the culture, the relative expression of DIgR2 mRNA in the cells cultured with LPS, IL-10, and CD40L was also determined in triplicate. DIgR2 levels were normalized to the cells of the third-day culture. Error bars indicate SE. (D) DIgR2 is a cell-surface monomeric glycoprotein. Lysates of DIgR2-transfected NIH/3T3 cells were treated in nonreducing or reducing conditions (left) or with N-glycosidase F in reducing conditions (right) and were then subjected to SDS-PAGE and blotted with anti-DIgR2 polyclonal antibody. (E) DIgR2 is expressed as a transmembrane protein. NIH/3T3 cells transfected with DIgR2 expression vector were stained with anti-DIgR2 polyclonal antibody (i) or purified rabbit serum (ii); NIH/3T3 cells transfected with CD54 were stained with anti-CD54 mAb (iii), and NIH/3T3 cells transfected with pcDNA3.1 were stained with anti-DIgR2 polyclonal antibody (iv). All of these transfectants were then counter stained with FITC-conjugated goat antirabbit or goat antimouse antibody and analyzed by fluorescence confocal microscopy. Data shown are representative of 2 independent experiments. Images were visualized using a Leica DMIRE2 microscope and a Leica 506140 40×/0.85 numeric aperture objective. Images were captured using a Leica DMIRE2 camera and Leica confocal software version 2.61.