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Not for use in diagnostic or therapeutic procedures. Not for resale.
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Principle of the Test
Six bead populations with distinct fluorescence intensities have been coated with
capture antibodies specific for IL-6, IL-10, MCP-1, IFN-γ, TNF, and IL-12p70
proteins. The six bead populations are mixed together to form the BD™ CBA which
is resolved in the FL3 channel of a flow cytometer such as the BD FACScan™
or BD FACSCalibur™ flow cytometer.
IL-12p70
TNF
IFN-γ
MCP-1
IL-10
IL-6
Figure 1
The capture beads, PE-conjugated detection antibodies, and recombinant standards
or test samples are incubated together to form sandwich complexes. Following
acquisition of sample data using the flow cytometer, the sample results are generated
in graphical and tabular format using the BD™ CBA Analysis Software. The
kit provides sufficient reagents for the quantitative analysis of 50 test samples
and the generation of two standard curve sets.
Advantages
The BD™ CBA provides several advantages when compared with conventional
ELISA methodology:
• The required sample volume is approximately one-sixth the quantity necessary
for conventional ELISA assays due to the detection of six analytes in
a single sample.
• A single set of diluted standards is used to generate a standard curve for
each analyte.
• A BD CBA experiment takes less time than a single ELISA and provides
results that would normally require six conventional ELISAs.
Limitations
The limit of detection of the BD™ CBA Mouse Inflammation Kit is comparable to
conventional ELISA, but due to the complexity and kinetics of this multi-analyte
assay, the actual limit of detection in a given experiment may vary slightly. Note
the reduced sensitivity of the Mouse MCP-1 assay (see Limit of Detection and
Precision information on pgs. 22 and 26 respectively).
The BD CBA is not recommended for use on stream-in-air instruments where
signal intensities may be reduced, adversely effecting assay sensitivity. Stream-in-
air instruments include the BD FACStar™ Plus and BD FACSVantage™
(BD Biosciences Immunocytometry Systems, San Jose, CA) flow cytometers.
Serum spike recoveries for IL-10 and TNF are lower than for the other proteins
in this assay. This variation is due to assay conditions and serum proteins. It may
affect quantitation of these proteins in serum samples.
Quantitative results or protein levels for the same sample or recombinant protein
run in ELISA and BD CBA assays may differ. A spike recovery assay can be
performed using an ELISA standard followed by BD CBA analysis to assess
possible differences in quantitation.
