Luminex 100 IS Developer Workbench Guide Version 2.3 - Manual (Page 63)

PN 89-00002-00-084 Rev. B 57
x
MAP Technology Oligonucleotide Coupling Protocol
Procedure
1. Disperse the pellet with sonication, and vortex the container for
20 seconds.
2. Transfer 5.0
× 10 6
of the stock microspheres into a 1.5 mL
microcentrifuge tube (See Technical note 2).
3. Microcentrifuge the xMAP microspheres at
≥ 8,000 × g for 1 to
2 minutes.
4. Aspirate the supernatant, being careful not to disturb the pellet.
Resuspend the microspheres in 50 µL of 0.1 M MES, pH 4.5.
Vortex and sonicate.
5. Prepare a 1:10 dilution of the 1 mM capture oligo in ddH
2
O. Add
2 µL (0.2 nanomole) of the 1:10 diluted capture oligo to the
resuspended microspheres. Vortex briefly (See Technical
note 3).
6. Immediately before use, prepare a 10 mg/mL solution of EDC
powder (See Technical note 4). Vortex until dissolved.
7. Add 2.5 µL of the fresh EDC solution to the xMAP
microspheres. Vortex immediately.
8. Incubate for 30 minutes at room temperature in the dark.
9. Repeat steps 6-8 with fresh EDC (for a total of two EDC
additions).
10. Add 1.0 mL of Tween 20 (0.02% v/v). Vortex.
11. Microcentrifuge xMAP microspheres at
≥ 8,000 × g for 1 to 2
minutes.
12. Aspirate the supernatant, being careful not to disturb the pellet.
13. Add 1.0 mL of SDS (0.1% w/v). Vortex.
14. Microcentrifuge the xMAP microspheres at
≥ 8,000 × g for 1 to
2 minutes.
15. Remove the supernatant, being careful not to disturb the pellet.
16. Resuspend the xMAP microspheres in 100 µL of TE Buffer
(10mM Tris, 1mM EDTA, pH 8.0). Vortex and sonicate for about
20 seconds.