1382-1406 Purification

1382-1406 purification on 10-30-2012 by Xiuli Transfection and cell viability: Paul did the transfection directly in the shaking flasks. The cells had very poor viability (shake didn't run properly) when collected.  Xiuli did transfections in the plate, cells started recovery on 3 rd day from transfection (didn't change into serum free medium, cells contaminated that week) and  stayed in the serum medim until collection.

Collect 30 ml Serum-free medium from each flasks (5) and and  plates (5) 20 ml and centriguged at 12000rpm for 15 mins.

Supernants:

Incubate supernanats (Paul's sample ) with protein A beads, washed with PBS and elute with 0.2 mM Glycine (pH=2.0), elution volume: 900 ul

Pellet:

Lysed pellet with 0.1% TX114 in PBS and collected supernant at 12000 rpm for 20 mins. Incubated the supernatants with protein A beads. Washed with PBS and eluted with Glycine (pH=2.0) for 4 times.

Load 20-35 ul on the 4-20% SDS gel

Run Paul's Sup and lysate samples, no protein show on the gel, Run Xiuli's samples from lysate.

IgG Pull down seems right, Lane 14 for 1383 have a faint band, can be the right size.