Bio-Rad Bio-Plex Pro™ Human Chemokine Assays - Manual (Page 20)

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8. Run Assay
Considerations
n
Bring all assay components and samples to room temperature
before use
n
Use calibrated pipets and pipet carefully, avoiding bubbles
n
Pay close attention to vortexing, shaking, and incubation instructions.
Deviation from the protocol may result in low assay signal and
assay variability
n
Assay incubations are carried out on a shaker at 850 ± 50 rpm at room
temperature (RT). Cover the plate with sealing tape and protect from
light with aluminum foil
Table 8. Summary of wash options and protocols. After each assay step, select the
appropriate Bio-Plex Pro ™ wash station program or perform the appropriate manual wash
step as summarized below.
Considerations When Using a Vacuum Manifold
n
After each incubation, place the filter plate on a calibrated vacuum
apparatus and remove the liquid by vacuum filtration
n
To wash, add 100 μl wash buffer to each well and remove the liquid as
before. Ensure that all wells are exposed to the vacuum
n
Thoroughly blot the bottom of the filter plate with a clean paper towel
between each vacuum step to prevent cross contamination
n
Place the assay plate on the plastic plate holder/tray as needed
n
Before each incubation, gently cover the plate with a new sheet of
sealing tape. Avoid pressing down on the wells to prevent leaking from
the bottom
Bio-Plex Pro or Bio-Plex Pro II Handheld Magnet or
Pro II Wash Station Wash Station Vacuum Manifold
Assay Step Magnetic Program Vacuum Program Manual Wash Steps
Add beads to plate MAG x2 VAC x2 2 x 100 μl
Sample incubation
Detection Ab incubation
MAG x3 VAC x3 3 x 100 μl
SA-PE incubation