Bio-Rad Bio-Plex Pro™ Rat Cytokine, Chemokine, and Growth Factor Assays - Manual (Page 28)

26
8. Cover plate with a new sheet of sealing tape and protect from light
with aluminum foil. Incubate on shaker at 850 ± 50 rpm for 30 min at
room temperature.
Prepare and Add Streptavidin-PE (SA-PE)
1. While the detection antibodies are incubating, use Table 23 or the
Calculation Worksheet on page 41 to calculate the volume of SA-PE
(100x) and assay buffer needed. Streptavidin-PE should be prepared
10 min before use.
2. Add the required volume of assay buffer to a 15 ml polypropylene
tube.
3. Vortex the 100x SA-PE for 5 sec at medium speed.
Perform a 30 sec spin to collect the entire volume at the bottom of
the tube.
4. Dilute SA-PE to 1x by pipetting the required volume into the 15 ml
tube. Vortex and protect from light until ready to use.
Each well of the assay requires 0.5 µl (100x stock) adjusted to a final
volume of 50 µl in assay buffer.
Table 23. Preparing 1x SA-PE from 100x stock (includes 25% excess volume).
5. After the detection antibody incubation, slowly remove and discard
the sealing tape.
6. Wash the plate three times with 100 µl wash buffer.
7. Vortex the diluted (1x) SA-PE at medium speed for 5 sec. Pour
into a reagent reservoir and transfer 50 µl to each well using a
multichannel pipet.
# of Wells 100x SA-PE, µl Assay Buffer, µl Total Volume, µl
96 60 5,940 6,000
48 30 2,970 3,000