Instructions for Lab Report 2:
Steps
1.) Take the memrane out of cassette. Put the protein side up (side that was touching the gel) into a small container. Then La Ponceau Stain with the help of Dr. Robson.
2.) Cover the membrane with blocking buffer. Then incubate at 4 degrees celcius overnight or on the rocker at room temperature for 3 to 4 hours.
3.) Dump out the blocking buffer down the sink. Then pour the entire contents of 1% Antibody tube over the membrane (in the fridge) and incubate overnight or on the rocker at room temperature for 1 to 2 hours.
4.) Wash 5x with wash buffer (cover the membrane with wash buffer and put on the rocker for 3 minutes and then dump the wash buffer down the sink and repeat.)
5.) Pour the entire contents of 2% antibody tube over the membrane(in fridge). Then incubate for 1 to 2 hours at room temperature on the rocker.
6.) Wash 5x with buffer as per(4). Pour last wash buffer in plastic baggie and store it in a zipped baggie that is immersed in the buffer in the fridge.
NEVER let the membrane dry out! (on steps 3 and 5 move the membrane to a smaller container if necessary to cover entirely with the antibody solution)
Steps 2/24/09
1.) Move the membrane back to the container PROTEIN side up.
2.) Wash 3X with TBS (tris-buffered saline). 3min/wash
3.) Dump the last wash down the sink; then cover with color developer solution and incubate on the rocker. (till the color develops)
