Code | CSB-PA010380PA25nme1HU |
Size | US$175 |
Order now | |
Image |
|
Have Questions? | Leave a Message or Start an on-line Chat |
Application | Recommended Dilution |
---|---|
WB | 1:500-1:2000 |
IHC | 1:20-1:200 |
IF | 1:50-1:200 |
Applications : Western Blot (WB), Immunohistochemistry (IHC)
Sample type: Human embryonic stem cell lines and HEK293T cells
Sample dilution: IHC: 1:200, WB: 1:1000
Review: IHC Method Human embryonic stem cell line, H7 cells were plated on matrigel pre-coated plate. 3 days after plating cells were fixed with 4% formaldehyde/PBS solution with 4% formaldehyde/PBS for 15 minutes, washed three times with PBS, and then permeabilized with 0.1 % Triton X-100/PBS. Fixed cells were subjected to blocking with 3% BSA/PBS solution and then incubated with primary antibody at 4°C overnight. Primary signals were detected using Alexa Fluor 488 and subsequently DAPI stained. IHC Result IHC result shows strong and clear H1.4 signals in nucleolus. W.B. Method HEKF 293T cells were plated on 6 well plate. 1day later cells were transfected with 1ug of control plasmid (pFUW-vector, addgene #14882) or pFUW-Flag ad Myc tagged-H1.4 plasmid. 2days additional incubation proteins were extracted using GST IP buffer (25 mM Tris-HCl, pH8.0, 150 mM NaCl, 100 mM KCl, 2 mM EDTA, 1% NP-40, and proteinase inhibitor cocktail) for 15 minutes on ice. Total protein was separated on a 12% PAGE gel and 5% skim milk were used for blocking. Primary antibodies were used in blocking solution at 4°C overnight. HRP-conjugated secondary antibodies were used for the detection. W.B. Results CSB-PA010380PA25 recognizes endogenous H1.4 proteins and ectopic overexpressed H1.4 proteins without any non-specific signals.
By Anonymous