Author: Steven Henikoff; Jorja G. Henikoff
Title: Profiling the epigenome at home Document date: 2020_4_17
ID: mojf4l6b_25
Snippet: • Resuspend cells in 50 µL Antibody buffer then 0.5 µL antibody (1:100) with gentle vortexing. Tip: For bulk processing, resuspend in Antibody buffer containing antibody (1:100) with gentle vortexing. Tip: We use 1:100 by default or the manufacturer's recommended concentration for immunofluorescence. • Place on a Rotator at room temperature and incubate 1-2 hr. In the lab, a nuclei prep was split and prepared as either native (Nat) or cross.....
Document: • Resuspend cells in 50 µL Antibody buffer then 0.5 µL antibody (1:100) with gentle vortexing. Tip: For bulk processing, resuspend in Antibody buffer containing antibody (1:100) with gentle vortexing. Tip: We use 1:100 by default or the manufacturer's recommended concentration for immunofluorescence. • Place on a Rotator at room temperature and incubate 1-2 hr. In the lab, a nuclei prep was split and prepared as either native (Nat) or cross-linked (XL), then aliquoted and frozen. At home, aliquots were thawed and libraries were prepared from 50,000 starting cells using this protocol with the following rabbit antibodies: H3K4me1-Th (Thermo #710795 lot 1998633); H3K4me1-Ep (Epicypher 13-0026 lot 28344001); H3K4me2-Mi (Millipore 07-030 lot 3229364); H3K4me2-Ep (Epicypher 13-0027); H3K4me3-Ac (Active Motif 39159 lot 22118006); H3K36me3-Ep (Epicypher Rabbit monoclonal #13-0031, lot 18344001); H3K9me3-Ab (Abcam ab8898 lot GR3302452-1); H3K27me3-Cs (CST #9733). The Tapestation image for 1/10th of each library is shown, where Native/XL is the molar ratio of yields over a 175-1000 bp range.
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