Patient P11 enriched_monoytes from day D15
PBMCs were isolated from the blood of healthy individuals or AD patients by the Ficoll gradient method. Blood was diluted with PBS at 1:1 ratio, layered on top of Ficoll-Paque Plus (Cytiva, catalog number 17144003) and centrifuged at room temperature for 20 minutes at 800 x g. The interface layer containing PBMCs was collected and washed twice with HBSS buffer (Cytiva, catalogue number SH30030.02). Then, cells were counted using the Cellometer Auto2000 Cell counter, and resuspended in freezing medium (90% heat inactivated FBS and 10% DMSO) at a concentration of 1-2x10^7 cells/ml. Cryovials were transferred at -80 C for 24 hours and then stored in liquid nitrogen at -140 C until the day of the experiment. The single-cell cDNA libraries were constructed using the Chromium Next GEM Single Cell 3' Reagent Kits v3.1 (10x Genomics, catalogue number 1000268) following the manufacturer's user guide. PBMC samples from patients P12, P13, P15, and P16 isolated on D0 and D15 and samples from patients P4, P7 and P11 isolated on D15 were thawed at 37C. Half of the cells in each vial were enriched in monocytes using the Human Pan Monocyte Isolation kit. PBMCs and enriched monocytes were stained with eBioscience Fixable Viability dye eFluor 660 for 15 minutes at 4C. After two washes with FACS buffer, cells were FACS-sorted on a Cytek Aurora CS sorter (Cytek) to remove dead cells. Live PBMCs and enriched monocytes were counted manually utilizing a hemocytometer and cell concentration was adjusted at 1700 cells/l. Cells and gel beads were loaded to the Chromium Next GEM Chip G which run on a Chromium X controller. Reverse transcription reactions generated barcoded full-length cDNA followed by disruption of emulsions using the recovery agent and cDNA clean up. Bulk cDNA was further amplified and cleaned up. Indexed libraries were constructed, equimolarly pooled and sequenced on an Illumina NovaSeq S2 aiming at  20,000 read pairs per cell at the Broad Institute.