Bio-Rad Bio-Plex Pro™ Magnetic Cell Signaling Assays - Manual (Page 32)

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Table 13. Troubleshooting guide (continued).
Problem Possible Causes Possible Solutions
High background Prolonged incubation of Follow the procedure incubation
detection antibodies and/or time precisely
streptavidin-PE
Wash steps performed Perform washes as described in
incorrectly or insufficient the assay instructions
washing volume
High assay CV Bottom of filter plate not dry Dry the bottom of the filter plate
with absorbent paper towel
(preferably lint-free) to prevent
cross-well contamination
Contamination with wash Be careful not to splash wash buffer
buffer during wash steps from well to well. Filter wells
completely to remove residual
buffer if using filter plate. Reduce
microplate shaking speed to
minimize splashing
Cell debris in lysate not cleared Centrifuge at 15,000 x g for 10 min
at 4°C to remove cellular debris
Shaking speed too high during Fully resuspend bead mixture at
assay incubation 900–1,100 rpm for 30 sec, then turn
down to 300–450 rpm for the
specified incubation time
Reagents and assay Bring all reagents and assay
components not equilibrated components to room temperature
to room temperature prior to dispensing