Bio-Rad Bio-Plex Pro™ TGF-β Assays - Manual (Page 3)

Running the Assay
Note: Make sure all assay components are at RT before proceeding.
1. Prewet filter plate with 100 µl Bio-Plex assay buffer (skip for flat bottom).
2. Vortex the diluted (1x) beads. Add 50 µl to each well of the assay plate.
3. Wash two times with 100 µl Bio-Plex wash buffer.
4. Vortex samples, standards, blank. Add 50 µl to each well.
5. Cover plate with sealing tape and protect from light with aluminum foil.
Incubate on shaker at 850 ± 50 rpm for 2 hr at RT.
Note: 850 rpm provides equivalent performance to recommended shaker
settings in previous manuals (1,100 rpm for 30 sec, 300 rpm for incubation).
6. With 10 min left in the incubation, vortex the 20x detection antibodies
for 5 sec and quick-spin to collect liquid. Dilute to 1x in detection
antibody diluent as shown below.
7. Wash the plate three times with 100 µl wash buffer.
8. Vortex the diluted (1x) detection antibodies. Add 25 µl to each well.
9. Cover and incubate at 850 ± 50 rpm, as in Step 5, for 1 hr at RT.
Meanwhile, prepare Bio-Plex Manager software protocol; enter
standard S1 values provided in the assay kit.
10. With 10 min left in the incubation, vortex the 100x SA-PE for 5 sec, quick-
spin to collect liquid, and dilute to 1x as shown below. Protect from light.
11. Wash the plate three times with 100 µl wash buffer.
12. Vortex the diluted (1x) SA-PE. Add 50 µl to each well.
# 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 5
# of Wells 100x SA-PE, µl Assay Buffer, µl Total Volume, µl
96 60 5,940 6,000