Clone K1 encodes a protein that is recognized by CD109 mAbs.

(A) By in vitro transcription–translation in the presence of³⁵S-methionine, followed by immunoprecipitation, CD109 clone K1 is shown to encode an approximately 160-kd protein (arrow, lane 1) that is recognized by mAb 1B3, but not by control antibodies, including CD71 mAb D51 (lane 2). Positions of radiolabeled molecular weight markers are shown on the left. Similar results were obtained using CD109 mAbs 8A3 and LDA₁ (not shown). (B) Expression of clone K1 confers CD109 mAb binding to CHO cells. Control mock-transfected CHO cells (top row), CHO cells transfected with empty pIRES-EYFP vector (middle row), and CHO cells transfected with CD109 expression vector pK1/YFP (bottom row) were stained with CD109 mAb 7D1-PE and 7-AAD. Viable (7-AAD⁻) cells were then gated (region R1), and single cells within this gate were identified by light scatter (R2, not shown). YFP-expressing cells within this population (R3, left column) were then analyzed for CD109 expression (R4, right column). Expression of the K1 cDNA confers high-level 7D1 binding to CHO cells. Similar specific staining was observed using phycoerythrin conjugates of CD109 mAbs 8A3 and TEA 2/16 (not shown), and with Gov^b antiserum.53 In contrast, staining was not detectable on control CHO cells or on CHO cells transfected with pIRES-EYFP–based vectors expressing K1 antisense or an irrelevant cDNA and was abrogated by treatment of K1-transfected cells with PI-PLC (not shown).