Chunk #74 — STAR Methods — Method Details — Immunocytochemistry and Immunohistochemistry

Source

iPSC-Derived Human Microglia-like Cells to Study Neurological Diseases.

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yes

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Alexa Fluor® conjugated secondary antibodies at 1:400 for 1 h at room temperature with slight shaking in the dark. After secondary staining, sections were washed in PBS 3 times for 5 min and mounted on glass slides. After mounting, slides were cover slipped with DAPI-counterstain mounting media (Fluoromount, southern Biotech). Primary antibodies: mouse anti-β3Tubulin (1:500; BioLegend, 801201)mouse anti-human Cytoplasm (SC121,1:100; Takara Bio Inc., Y40410),mouse anti-human Nuclei (ku80, 1:100; Abcam, ab79220)chicken anti-GFAP (1:500; Abcam, ab4674)rabbit anti-Iba1 (1:500; Wako; 019-19741)goat anti-Iba1 (1:100; Abcam ab5076) * recommend use with Alexa Fluor 488 or 555 secondary antibody only.mouse anti-ITGB5 (1:500; Abcam, ab177004)mouse anti-MMP-9 (1:500; EMD Millipore, AB19016)mouse anti-human Mertk (1:500; BioLegend, 367602)rabbit anti-P2ry12 (1:125; Sigma; HPA014518)rabbit anti-Pros (1:500; Abcam, ab97387)rabbit anti-PU.1 (1:500; Cell Signaling Technology, 2266S)rabbit anti-human Tmem119 (1:100; Abcam, ab185333)goat anti-human Trem2 (1:100; R&D Systems, AF1828)rabbit anti-Tgfbr1 (1:500; Abcam, ab31013).rabbit anti-Cx3cr1 (1:1000; Bio-Rad/AbD Serotec, AHP1589)