Nuclear Protein Kit

  
        

 

Product Cat. # Description Pack Price
Cell Nuc. extration SINP001a Native cell nuclear extraction (Manual). 50 samples $99
Tissue Nuc. extration SINP001b Native tissue nuclear extraction (Manual). 50 samples $99

A. Introduction:

Increased awareness of cellular proteins as potential drug targets or as biological markers for the detection of disease conditions necessitates the development of new protein sample preparation techniques from tissue or cells. 

This nuclear extraction kit covers 1) the extraction of nuclear proteins from whole cells/tissues, 2) removal of interfering substances, and 3) prevention of protein degradation, providing nuclear proteins that are directly compatible with downstream analytical applications. 

Viagene Nuclear extraction kits provide reproducible, reliable and convenient protein sample preparation to achieve biologically relevant data by EMSA, immunoblotting, ELISA, array, activity assay, 2D gel electrophoresis and mass spectrometry. 
  

B. Major procedure:NFkB EMSA

1. Touch cells - nuclear protein extraction:

1. aspirate medium
2. wash with ice-cold PBS
3. add 3 mL ice-cold PBS and scrape cells gently into 15mL tube
4. centrifuge 5 minutes at 500 rpm, 4OC
5. carefully aspirate supernatant with pipet
6. resuspend pellet in 1 mL ice-cold Working Sol. I
7. incubate 15 minutes on ice to allow cells to swell
8. add 100 ul of Solution II to 1 mL  ice-cold Working Sol. I
9. vortex 10 seconds
10. centrifuge 2-3 minutes at maximum speed (~15K RPM)
11. carefully aspirate supernatant with pipet; this is the cytoplasmic fraction
12. resuspend the pellet in 175 ul ice cold Working Sol. III
13. vortex 30 seconds; rotate vigorously at 4OC 30 minutes
14. centrifuge 15 minutes at maximum speed; remove the supernatant to a fresh, chilled tube. leave on ice if using immediately, or store aliquots at –80OC until use. DO NOT freeze/thaw.
15. assay for protein concentration (Pierce’s BCA kit).

2. Suspension cells - nuclear protein extraction: (see other section)
  

C. Pack (for 50 samples) list:

  Components Unit Pack
5 X Buffer A 1 bottle 35 ml/bottle
2X Buffer B 1 vial 1.5 ml/vial
Solution I 1 vial 1.75 ml/vial
Solution II 1 vial 1.75 ml/vial
Solution III 1 vial 1.5 ml/vial
PMSF Solution 1 vial 0.85 ml/vial
User Manual 1 set 1

 

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