Bio-Rad Bio-Plex Pro™ Magnetic Cell Signaling Assays - Manual (Page 3)

2. Prime wash station or prepare handheld magnetic washer. (Section 3)
3. Calibrate the Bio-Plex system by following the prompts within
Bio-Plex Manager ™ software. (Section 5)
4. With 10 min left in the overnight incubation, vortex detection antibodies
for 5 sec and quick-spin to collect liquid. Dilute to 1x in detection
antibody diluent as shown below.
5. Wash the plate three times with 200 μl wash buffer.
6. Vortex the diluted (1x) detection antibodies. Add 25 μl to each well.
7. Cover and incubate in the dark for 30 min at RT. Shake at
900–1,100 rpm for 30 sec and then turn down to 300–450 rpm for
the specified incubation time.
Meanwhile, prepare the Bio-Plex Manager software protocol. (Section 5)
8. With 10 min left, vortex streptavidin-PE (SA-PE) for 5 sec and quick-
spin to collect liquid. Dilute to 1x as shown below and protect from light.
9. Wash the plate three times with 200 μl wash buffer.
10. Vortex the diluted (1x) SA-PE. Add 50 μl to each well.
11. Cover and incubate in the dark for 10 min at RT. Shake at
900–1,100 rpm for 30 sec and then turn down to 300–450 rpm for
the specified incubation time.
12. Wash the plate three times with 200 μl wash buffer.
# of Wells 20x Detection Ab, µl Detection Ab Diluent, µl Total Volume, µl
96 150 2,850 3,000
Bio-Plex Pro Assay Quick Guide 3
# of Wells 100x SA-PE, µl Detection Ab Diluent, µl Total Volume, µl
96 60 5,940 6,000