ProteinSimple eBook - 29

Simplifying Protein Analysis * Multi-Analyte, Automated, Microﬂuidic Immunoassay Platform

IL-1β Calibration Curves

IL-10 Calibration Curves

10,000
1,000

100

Signal (RFU/ms)

1,000

1.0

100
10
1.0

Ella

Ella
ELISA

ELISA

0.1

0.1
0

100

1,000

10,000

Concentration, pg/mL

IL-5 Calibration Curves

10,000

100

1,000

10,000

IL-12 Calibration Curves

10,000

SIMPLE PLEX VS. TRADITIONAL PLATE ELISA

1,000

Signal (RFU/ms)

1,000

Signal (RFU/ms)

Concentration, pg/mL

100
10
1.0

Ella

ELISA

0.1

0.1
0

100

Concentration, pg/mL

1,000

10,000

100

CV) for IL-6, and 100% (9.8% CV) for IL-10. Mean recovery for the 40 pg/mL
(pmol/L) point, made in calibration buffer, for the constant assays back-ﬁt
to each panels speciﬁc calibration curve was 104% (4% CV) for IL-1β, 101%
(5% CV) for IL-6, and 106% (7% CV) for IL-10. These data show that for the
immunoassays studied the accuracy and precision of an individual assay is not
negatively impacted by changing the immunoassay in neighboring channels
within the Simple Plex cartridge.)

1,000

10,000

Concentration, pg/mL

Figure 5. Simple Plex assay vs. traditional ELISA. Calibration curves graphed
with relative units.

Calibration curves run in the ELISA format showed dynamic ranges around
2 to 2.5 orders of magnitude, whereas in the Simple Plex assay format dynamic
ranges with the same standard were all greater than four orders of magnitude
showing a twofold increase over the ELISA format (Figure 5). Furthermore, the
increase in dynamic range of the Simple Plex assay format spanned both ends of
the curve showing improved sensitivity for measuring low concentrations and
expanded range for elevated samples. In particular, the increased sensitivity is
especially crucial for cytokines as most healthy levels range in the single digit
pg/mL range.
SIMPLE PLEX VS. TRADITIONAL MULTIPLEX ELISA

Calibration curves for IL-5 were run as a traditional Simple Plex assay format
and as a traditional multiplex with increased complexity of cocktailed dAbs
(Figure 6). The data revealed that as the number of dAbs in the cocktail increased,
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