Bio-Rad Bio-Plex Pro™ Magnetic Cell Signaling Assays - Manual (Page 19)

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7. Dilute coupled beads to 1x by pipetting the required volume into the
tube containing wash buffer. Vortex.
Each well of the assay requires 2.5 μl of the 20x stock adjusted to a
final volume of 50 μl in wash buffer.
Note: To minimize volume loss, use a 200–300 μl capacity pipet to
remove beads from the 20x stock tube. If necessary, perform the volume
transfer in 2 steps. Do not use a 1,000 μl capacity pipet and/or wide
bore pipet tip.
8. Protect the beads from light with aluminum foil. Equilibrate to room
temperature prior to use.
9. Cover unused wells of the assay plate with sealing tape.
10. Prewet the filter plate. Skip this step if using a flat bottom plate.
a. Prewet the wells with 200 μl wash buffer and remove the liquid by
vacuum filtration. Dry the bottom of the filter plate thoroughly by
blotting on a clean paper towel.
11. Vortex the diluted (1x) beads for 15 sec at medium speed. Transfer
50 µl to each well of the assay plate.
12. Wash the plate two times with 200 µl wash buffer according to your
method of choice.
# of Wells 20x Beads, µl Wash Buffer, µl Total Volume, µl
96 288 5,472 5,760
48 144 2,736 2,880
Preparing 1x coupled beads from 20x stock (includes 20% excess volume):
Table 6. Premixed panel or one singleplex assay.
20x Beads, µl 20x Beads, µl Wash
# of Wells Singleplex #1 Singleplex #2 Buffer, µl Total Volume, µl
96 288 288 5,184 5,760
48 144 144 2,592 2,880
Table 7. Mixing two singleplex assays.