Bio-Rad Bio-Plex Pro™ Human Cancer Biomarker Assays - Manual (Page 18)

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6. Prepare Coupled Beads
1. Use Tables 6–7 or the Calculation Worksheet on page 35 to calculate
the volume of coupled beads and assay buffer needed.
2. Add the required volume of Bio-Plex ® assay buffer to a 15 ml
polypropylene tube.
3. Vortex the 20x stock of coupled beads at mid speed for 30 sec.
Carefully open the cap and pipet any liquid trapped in the cap back
into the tube. This is important to ensure maximum bead recovery.
Do not centrifuge the vial; doing so will cause the beads to pellet.
4. Dilute coupled beads to 1x by pipetting the required volume into the
15 ml tube. Vortex.
Each well of the assay requires 2.5 μl of the 20x stock adjusted to a
final volume of 50 μl in assay buffer.
5. Protect the beads from light with aluminum foil. Equilibrate to room
temperature prior to use.
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 two steps. Do not use a 1000 μl capacity pipet and/or wide bore pipet tip.
Preparing 1x coupled beads from 20x stock (includes 20% excess volume)
Table 6. Premixed panel or one singleplex assay.
Table 7. Mixing singleplex assays.
# of Wells 20x Beads, µl Assay Buffer, µl Total Volume, µl
96 288 5,472 5,760
48 144 2,736 2,880
20x Beads, µl 20x Beads, µl
# of Wells Singleplex #1 Singleplex #2 Assay Buffer, µl Total Volume, µl
96 288 288 5,184 5,760
48 144 144 2,592 2,880