Journal: bioRxiv

Article Title: The MS4A gene cluster is a key regulator of soluble TREM2 and Alzheimer disease risk

doi: 10.1101/352179

Figure Lengend Snippet: (A) Levels of sTREM2 measured by ELISA in the cell media of human macrophages on day 7 in culture with or without stimulation with IL-4. (B) Effect on levels of sTREM2 in cell culture media by treatment with antibodies against MS4A4A and MS4A6A for 48 h. Two different anti-MS4A4 antibodies were tested, antibody A (Abcam) directed against the N-Terminal cytoplasmic domain or antibody B (Biolegend) directed against the extracellular domain of the protein. These analyses were done without or with IL-4 in culture. Bars are means ± SD. *p<0.05; **p<0.005; ***p<0.005. Pvalues were calculated by Mann-Whitney test or Kruskal Wallis H test for multigroup comparison. (C) Confocal images of human macrophages treated or not with IL-4 for 48 hours and stained for TREM2 (red) and MS4A4A (green) antibodies. Arrowheads (white) indicate colocalizing signal. Scale bar 10 μm.

Article Snippet: The assay consists of a Streptavidin-coated 96-well plates (MSD Streptavidin Gold Plates, cat. no. L15SA-1); a biotinylated polyclonal goat IgG anti-human TREM2 antibody (R&D Systems, cat. no. BAF1828; 0.25 μg/mL, 25 μL/well) as capture antibody, which is raised against amino acids 19174 of human TREM2; a monoclonal mouse IgG anti-human TREM2 antibody (Santa Cruz Biotechnology, B-3, cat. no. sc373828; 1 μg/mL, 50 μL/well) as a detection antibody, which is raised against amino acids 1-160 of human TREM2; and a SULFO-TAG-labeled goat polyclonal anti-mouse IgG secondary antibody (MSD, cat. no. R32AC; 0.5 μg/mL, 25 μL/well).

Techniques: Enzyme-linked Immunosorbent Assay, Cell Culture, MANN-WHITNEY, Staining