The other reasons being the team comprising of director Prajith, scenarist Sajeev Pazhoor and Biju ettan
After rehydration, samples were focused using an Ettan
IPGphor IEF unit (GE healthcare) according to the manufacturer's conditions.
After the fire destroyed their residence, the Louds moved to a new home on a small island near the west end of Van Ettan
Lake, some seven miles northwest of Oscoda.
For the first dimension, IEF was performed using the Ettan
IPGphor Isoelectric Focusing System (General Electric Healthcare, Piscataway, NJ, USA) while the SDS-PAGE was done using the gel caster system and the Mini-protean II (Bio-Rad, Hercules, CA, USA).
After the strips were subjected to isoelectric focussing, the IPG strip was equilibrated in solution (6 mol/L urea, 2 mol/L thiourea, 2% CHAPS, 0.5% IPG buffer containing 1% DTT w/v for 15 min, then incubated in 4% iodoacetamide w/v for 15 min on Ettan
IPGphor Isoelectric Focusing System (GE Amersham).
The first dimension isoelectric focusing (IEF) was carried out in an Ettan
IPGphor II Manifold applying a manufacturer's standard protocol with some modifications (strips were pre-focused at 500 V for 1 hour, 1,000 V for 1 hour and 8,000 V for 7 hours).
The first dimension separation (or isoelectric focusing) was performed by the Ettan
IPGphor IIIIEF System (GE healthcare) at 20[degrees] C in a stepwise voltage increase to reach 8,333 Vhrs with the limited of 50 [micro]A/strip.
The IEF in the Ettan
IPGphor 3 (GE Healthcare) system was carried out according to the manufacturer's manual.
Isoelectric focusing was done with an Ettan
IPGphor3 (GE) and maintained at 20[degrees]C.
Isoelectric focusing was run up to 90,000 Vh (Ettan
II IPGphor system, GE Healthcare).
After an overnight passive rehydration at room temperature, proteins were focused on an Ettan
IPGphor II (Amersham Biosciences) until reaching 24,000 total voltage hours (VhT).