Luminex 100 User Manual Version 1.7 - Manual (Page 121)

x MAP Technology Protocols
PN 89-00002-00-063 Rev. A 9 - 11
• REACTION BUFFER:
Phosphate Buffered Saline (pH 7.3 ± 0.1), Bovine Serum
Albumin (10 mg/ml)
• BLOCKING/STORAGE BUFFER:
Phosphate Buffered Saline (pH 7.3 ± 0.1), Tween 20
(0.02% v/v), Bovine Serum Albumin (1 mg/ml), Sodium Azide
(0.05% w/v)
Preparation
1. Dilute the Biotin-conjugated molecule in REACTION BUFFER
to a concentration of 4 - 4000 nM (See Technical note 1).
Procedure
1. Centrifuge the Avidin-modified xMAP microsphere stock for 1
minute at 8,000 × g or higher.
2. Disperse the pellet with sonication, and vortex the container for
20 seconds.
3. Dispense 1.0 × 10 5 Avidin-modified xMAP microspheres into a
1.5 ml microcentrifuge tube.
4. Centrifuge the xMAP microspheres for 1 minute at 8,000 × g or
higher and remove the supernatant.
5. Add 250 µl of the REACTION BUFFER (See Technical
note 2). Vortex.
6. Add 250 µl of the Biotin-conjugated molecule solution to the
xMAP microsphere suspension. Vortex immediately.
7. Rotate the mixture for 30 minutes in the dark at room
temperature.
8. Centrifuge the microspheres for 1 minute at 8,000 × g or higher.
Aspirate the supernatant.
9. Add 500 µl of BLOCKING/STORAGE BUFFER to the
microspheres and vortex. Centrifuge the tube for 60 seconds.
Aspirate the supernatant.
10. Repeat step 9.
11. Resuspend the microspheres in the desired volume of
BLOCKING/STORAGE BUFFER.
12. Enumerate the microsphere preparation.
13. Store the preparation at 2
o
C - 8°C. Protect it from light.