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1. For confocal laser microscope observation, Isolation cells were plated on a chamber slide with MEF feeders.3 X+ d3 F, E+ E0 u
2. Cells were washed twice in PBS and fixed with 4% paraformaldehyde in PBS for 15 min at room temperature.- v: _- c7 [8 N% m2 r7 f% I) i: m
3. Fixed cells were permeabilized with 0.5% Triton X-100 in PBS for 10 min. : b, ^! M% n% B3 z& s$ \# K; R$ D
步骤3后 要不要用PBS 清洗后进行以下步骤? P0 A8 g& G: m# R0 C. L
4. Then cells were blocked with 5% bovine serum albumin in PBS for 1 h. 9 E2 J# W6 |9 l9 |* X2 W7 p( E
5. The cells were incubated with primary antibodies for 16 h at 4 ° C. (过夜?)
; H, r+ ?" q4 z( ^( Z/ w& E- S) g 6. The Cells were washed twice in PBS 。1 J( h, {' x% j# m. u9 n7 d' i
7. Then the cells were incubated with secondary antibodies for 1 h at 37 ° C.5 J6 v1 _9 t4 W5 U: D$ E
8. For nuclear staining, the cells were stained with Hoechst 33,258 for 15 min at room temperature.
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4 {- w4 K: X- h8 R4 G* K请大侠 批评改正 |
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