Bio-Rad Human MMP and TIMP Assays - Manual (Page 3)

Running the Assay
Note: Make sure all assay components are at RT before pipetting. Vortex at
medium speed.
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 the plate two times with 100 µl Bio-Plex wash buffer.
4. Vortex samples, standards, blank, and control. 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 at RT for 1 hr.
6. With 10 min left in the incubation, vortex detection antibodies for 15 sec
and quick-spin to collect liquid. Dilute to 1x 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 described above, in the
dark for 30 min at RT. Meanwhile, prepare Bio-Plex Manager software
protocol; enter standard S1 values and units provided in the assay kit.
10. With 10 min left in the incubation, vortex 100x SA-PE for 5 sec and
quick-spin to collect liquid. Dilute to 1x as shown below and 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.
13. Cover and incubate at 850 ± 50 rpm, as described above, in the dark
for 10 min at RT.
14. Wash the plate three times with 100 µl wash buffer.
# of Wells 20x Detection Ab, µl Detection Ab Diluent HB, µl Total Volume, µl
96 150 2,850 3,000
Bio-Plex Pro Human MMP Assays Quick Guide
# of Wells 100x SA-PE, µl Assay Buffer, µl Total Volume, µl
96 60 5,940 6,000