Bio-Rad Nonmagnetic Beads and Related Reagents - Manual (Page 18)

22. Place the tube into a magnetic separator and allow separation to
occur for 30 to 60 sec.
23. With the tube still positioned in the magnetic separator, remove the
supernatant. Take care not to disturb the beads.
24. Remove the tube from the magnetic separator and resuspend the
coupled beads in 500 µL of PBS, pH 7.4.
25. Place the tube into a magnetic separator and allow separation to
occur for 30 to 60 sec.
26. Resuspend the coupled beads with 250 µL of blocking buffer.
27. Vortex the beads at medium speed for 15 sec.
28. Cover the coupling reaction tube with aluminum foil and agitate the
beads on a shaker (or rotator) for 30 min at room temperature.
29. Place the tube into a magnetic separator and allow separation to
occur for 30 to 60 sec.
30. With the tube still positioned in the magnetic separator, remove the
supernatant. Take care not to disturb the beads.
31. Remove the tube from the magnetic separator and resuspend the
beads in 500 µL of storage buffer by vortex 20 sec.
32. Place the tube into a magnetic separator and allow separation to
occur for 30 to 60 sec.
33. Remove the tube from the magnetic separator and resuspend the
coupled and washed beads in 150 µL of storage buffer.
34. Determine the bead concentration using a Coulter Z2 counter or a
hemocytometer.
35. Store coupled beads refrigerated at 2–8°C in the dark.
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