NIH 3T3 Nuclear Cell Lysate

Mouse

Immunoprecipitation, Western Blot

Use it directly for immuno-precipitation, or heat lysate with SDS gel loading buffer to 95C for 5 minutes followed by rapid cooling for western blot application. If dissociating conditions are required, add reducing agent prior to heating.

NIH/3T3 (mouse NIH/Swiss embryo) nuclear extract lysate (non-denatured)

Embryo, contact-inhibited

Cell

Embryo

Native

Nuclear

Nuclear extract was prepared by using a modified protocol of Dignam et al. Cells were Harvested and homogenized in Buffer A, and then centrifugated at 25,000 g for 20 minutes to remove cytoplasm and pellet the nuclei. The pellet was re-suspended in Buffer C, and then the suspensions were centrifuged to collect nuclear extract. The supernatant was dialyzed against Buffer D. The dialysate was then centrifuged, divided into aliquots, and stored at -80C. The protein concentration was determined by the method of Bradford. The lysate was adjusted to 2 mg/ml.

20mM HEPES pH 7.9, 20%(v/v) glycerol, 50mM KCl, 0.2 mM EDTA, 0.5 mM DTT & 0.5 mM PMSF.

2 mg/ml

The product is shipped with dry ice or equivalent. Upon receipt, store it immediately at the temperature recommended below.

Store at -80C. Avoid freeze-thaw cycles.