Biology Reference
In-Depth Information
Next day, aspirate wells manually using a very fine plastic tip placing on a single
position close to the side of the well bottom. It is important not to scratch
the surface in the center of the well.
Load 10
L blocking solution per well to columns 2-23.
Centrifuge 5 min at 500
m
g
.
Incubate 30 min at RT.
Aspirate wells.
Load 20
L purified water per well. (On epMotion, use multidispense, change
tips after command and water liquid-type settings.)
Aspirate wells.
Load 20
m
L purified water per well.
Aspirate wells and dry plate in a laminar flow (tissue culture) hood for 2 h.
Store plate in a plastic bag at 4
C (may be stored for a few weeks).
m
15.7.4
Step 4: immobilization of NeutrAvidin
15.7.4.1
Materials
NeutrAvidin (Pierce 31000)
15.7.4.2
Solutions and buffers
10 mg/mL NEUTRAVIDIN:
10 mg NeutrAvidin.
990
L purified water.
Aliquot (50
m
80
C.
L per vial) and store at
m
0.37% DM:
10
L 10% DM
m
260
L purified water
m
PRIMING SOLUTION:
10
L 0.37% DM
m
990
L PBS
m
NEUTRAVIDIN SOLUTION (0.06 mg/mL):
6
L 10 mg/mL NeutrAvidin
m
40
m
L 5% BSA.
10
m
L 0.37% DM.
944
L PBS.
Centrifuge 10 min at 12,000
m
g
.
BLOCKING SOLUTION:
40
m
L 5% BSA
10
L 0.37% DM
m
944
L PBS
m
