Bio-Rad Apoptosis Assays - Manual (Page 14)

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2. Bring the 10x assay buffer to room temperature (RT).
a. Mix by inversion to ensure all salts are in solution.
b. Prepare 1x assay buffer — dilute 1 part 10x assay buffer (60 ml) with
9 parts of dH
2
0 (540 ml).
Test Sample Preparation
Thaw and dilute samples within 1 hr of use. Remove any particulates by
centrifugation or filtration. Avoid multiple freeze and thaw cycles.
Dilution of Standard (1:3 Serial Dilution)
This preparation provides sufficient volume to run duplicate standard
dilution curves. Ensure that each new standard is mixed well by vortexing
before proceeding to the next dilution. Change tips between each dilution.
Note: The product data sheet in each kit lists the most concentrated
point on the standard curve (S1). Enter the values and units into
Bio-Plex Manager
™ software as instructed in the Read Plate section.
1. Label 8 polypropylene tubes S1 through S8. Label an additional
tube Blank.
2. Transfer the reconstituted standard into the tube labeled “S1.”
3. Add the appropriate amount of the standard diluent into the labeled
tubes according to Figure 3 (this will be sufficient for duplicate
standard curves and blanks).
4. Prepare working standards (S2–S8) by 1:3 (threefold) serial dilution.
Transfer the appropriate volume of standard into each of the labeled
tubes containing standard diluent as outlined above.
5. Vortex each standard at a medium setting for 5 sec before proceeding
with the next serial dilution. Change pipet tip at each dilution step.