Bio-Rad Bio-Plex Pro™ Rat Cytokine, Chemokine, and Growth Factor Assays - Manual (Page 26)

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Prepare and Add Detection Antibodies
Instructions are provided for diluting the detection antibodies to a 1x
concentration. When mixing diabetes and cytokine assays, keep in mind
the stock concentrations of detection antibodies as shown below.
Table 17. Stock concentration of detection antibodies.
1. While the samples are incubating, use Tables 18–22 or the
Calculation Worksheet on page 41 to calculate the volume of
detection antibodies and detection antibody diluent needed.
Detection antibodies should be prepared 10 min before use.
2. Add the required volume of Bio-Plex detection antibody diluent to a
15 ml polypropylene tube.
3. Vortex the stock detection antibodies for 15–20 sec at medium
speed, then perform a 30 sec spin to collect the entire volume at the
bottom of the tube.
4. Dilute detection antibodies to 1x by pipetting the required volume into
the 15 ml tube.
Each well of the assay requires either 1.25 μl (20x stock) or 2.5 µl (10x
stock) adjusted to a final volume of 25 μl in detection antibody diluent.
Preparing 1x detection antibodies from 10x stock (includes 25% excess volume)
Table 18. Premixed panel or one singleplex assay.
Stock Concentration of
Assay Detection Antibodies
Human and mouse cytokines (groups I, II) 10x
Mouse cytokines (group III) 20x
Rat cytokines (group I) 20x
Human, mouse, and rat diabetes 20x
10x Detection Detection Antibody
# of Wells Antibodies, µl Diluent, µl Total Volume, µl
96 300 2,700 3,000
48 150 1,350 1,500