Procedure – Luminex 100 User Manual Version 1.7 User Manual

Page 119

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MAP Technology

Protocols

PN 89-00002-00-063 Rev. A

9 - 9

Procedure

1. Centrifuge the xMAP microsphere stock for 1 minute at a

minimum of 8,000 x g.

2. Disperse the pellet with sonication, and vortex the container for

20 seconds.

3. Dispense 5.0

× 10

6

of the xMAP microspheres from the stock

tube into a 1.5 ml microcentrifuge tube (See Technical note 2).

4. Microcentrifuge the xMAP microspheres at 8,000

× g or higher

for 1 minute.

5. Remove the supernatant, being careful not to disturb the pellet.

6. Add 50 µl of 0.1 M MES (pH 4.5). Vortex and sonicate.

7. Add 0.2 nmole of amino-substituted oligonucleotide (i.e., 2 µl of

a 1:10 dilution of the 1 mM solution). Vortex briefly (See
Technical note 3).

8. Immediately before use, prepare a 10 mg/mL solution of EDC

powder (See Technical note 4). Vortex until dissolved.

9. Add 2.5 µL of the fresh EDC solution to the xMAP

microspheres. Vortex immediately.

10. Incubate for 30 minutes at room temperature in the dark.

11. Repeat steps 8-10 with fresh EDC (for a total of two EDC

additions).

12. Add 1.0 ml of Tween 20 (0.02% v/v). Vortex.

13. Centrifuge the xMAP microspheres at 8,000

× g or higher for 1

minute.

14. Remove the supernatant, being careful not to disturb the pellet.

15. Add 1.0 ml of SDS (0.1% w/v). Vortex.

16. Centrifuge the xMAP microspheres at 8,000

× g or higher for 1

minute.

17. Remove the supernatant, being careful not to disturb the pellet.

18. Resuspend the xMAP microspheres in 100 µl of TE (10mM Tris,

1mM EDTA, pH 8.0).

19. Enumerate the xMAP microsphere preparation.

20. Store the preparation at 2

o

C - 8°C. Protect it from light.

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