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1. For confocal laser microscope observation, Isolation cells were plated on a chamber slide with MEF feeders.
. \4 m8 I. U: J5 w* Z 2. Cells were washed twice in PBS and fixed with 4% paraformaldehyde in PBS for 15 min at room temperature.
& e7 K; o O9 B. _+ W: m$ R 3. Fixed cells were permeabilized with 0.5% Triton X-100 in PBS for 10 min.
& \4 r6 o0 o$ }% o& A步骤3后 要不要用PBS 清洗后进行以下步骤?
# D' p" |& [1 n% c& K) g/ j 4. Then cells were blocked with 5% bovine serum albumin in PBS for 1 h. 4 V) ]6 t0 ~$ X! u) K' C
5. The cells were incubated with primary antibodies for 16 h at 4 ° C. (过夜?)1 a d% V% b, b) K2 w
6. The Cells were washed twice in PBS 。" R2 o- c: V+ b" j6 Y1 q0 S; V
7. Then the cells were incubated with secondary antibodies for 1 h at 37 ° C.
( Q# h# J6 @: e 8. For nuclear staining, the cells were stained with Hoechst 33,258 for 15 min at room temperature.9 ]* j4 y8 I2 g# ?% H1 ?# v+ V2 o
& a; N+ j5 d8 m3 n6 g" x$ d请大侠 批评改正 |
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