Anti cd19 percp 1d3

MOG_42-55-IA^g7, hen egg lysozyme (HEL)_11-25-IA^g7, OVA_141-160-IA^g7, and human CLIP_87-101-IA^g7 monomers and tetramers were provided by the National Institute of Health Tetramer Core Facility at Emory University. Mononuclear cells were stained with tetramer immediately post-Percoll gradient isolation. Live cells were recovered from lymphocyte culture by subjecting cells to a Ficoll gradient (Mediatech) and then stained with 4μg/ml tetramer in culture medium for 4h at 37°C. Cells were then stained with anti-CD4-FITC (RM4-5, BD Pharmingen), anti-CD11b-PerCP (M1/70, BD Pharmingen), anti-CD11c-PerCP (HL3, BD Pharmingen), anti-CD19-PerCP (1D3, BD Pharmingen), anti-CD44-PE-Cy7 (IM7, Biolegend), and anti-CD8a-V450 (53-6.1, Tonbo Biosciences) for 30 minutes on ice. Flow cytometry was performed on either a BD Biosciences FACSVerse or LSRII flow cytometer. Data were analyzed using FloJo software (Tree Star).

Bone marrow (BM) transplantations were conducted as previously described (20 (link)). To distinguish donor from recipient BM-derived cells colonies of WT, ApoA-I^−/− and ApoA-I^tg mice expressing the differential Ly5.1 (CD45.1) alloantigen were derived by crossing with the B6.SJL-Ptprca Pep3b/BoyJ strain of C57BL/6J mice (Jackson Laboratories). Ly5.1 C57BL/6J, Ly5.1 ApoA-I^−/− and Ly5.1 ApoA-I^tg recipients were lethally irradiated (two doses of 450 RADS ionizing radiation spaced by three hours) and transplanted at six weeks of age with 5×10⁶ BM mononuclear cells from Ly5.2 WT or Ly5.2 Sle123 donor mice. Peripheral blood samples were collected by retro-orbital puncture and processed for flow cytometric analysis of reconstitution of major hematopoietic lineages using the following conjugated antibodies: anti-CD4 [RM4–5] PerCP, anti-CD8 [53–6.7] FITC, anti-CD11b [M1/70] PE, anti-CD19 PerCP [1D3] (BD Pharmingen, San Jose, CA), anti-Ly5.1 [A20] FITC or PE, anti-Ly5.2 [104] APC (Southern Biotech, Birmingham, AL). No aberrant ApoA-I specific antibody responses were generated as a result of BM transfer into WT or ApoA-I gene targeted mice (by anti-human ApoA-I and anti-mouse ApoA-I ELISA; as in (18 (link)); data not shown).