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THUNDER™ TR-FRET with BMG LABTECH’s PHERAstar® FSX: a powerful approach for high sensitivity cytokine quantification

2022-03-15

As part of our ongoing strategic collaboration with BMG LABTECH, we have recently evaluated the performance of a panel of five THUNDER™ TR-FRET human cytokine assays on the top-of-the-line PHERAstar^® FSX multi-mode microplate reader. The results, summarized in the Application Note 365, showed a superb assay performance of our assays on this HTS instrument.

Figure 1: THUNDER™ Human IL-1β standard curve.

Figure 2: A linear plot of the low concentration part of the THUNDER™ Human IL-1β standard curve.

Table 1: Performance of THUNDER™ human cytokine assays on the PHERAstar FSX and the competing HTS reader.

Analyte	Assay parameter	PHERAstar FSX	Competitor
IFNγ	LOD (pg/mL)	6	24
	LLOQ (pg/mL)	33	121
	S/B	15	9
IL-1β	LOD (pg/mL)	1	26
	LLOQ (pg/mL)	7	141
	S/B	23	17
IL-12	LOD (pg/mL)	3	20
	LLOQ (pg/mL)	19	98
	S/B	22	15
TNFα	LOD (pg/mL)	3	13
	LLOQ (pg/mL)	41	62
	S/B	57	45
CCL2	LOD (pg/mL)	3	13
	LLOQ (pg/mL)	17	61
	S/B	17	15

Highlights

The THUNDER™ human IFN-gamma, IL1-beta, IL-12, CCL2, and TNF-alpha assay kits were tested in a 384-well format using the standardized protocols.
Assays were read on the PHERAstar FSX equipped with a 377 nm laser excitation source.
For comparison, plates were also read on a competing HTS microplate reader equipped for TRF laser excitation.
The limit of detection (LOD) and lowest limit of quantification (LLOQ) were calculated by adding 2 or 10 standard deviations (SD), respectively, to the mean background counts (zero standard; 12 replicate wells).
For all assays tested, the PHERAstar FSX exhibited broader dynamic ranges and higher sensitivity compared to the competitor and our internal data, decreasing the LOD to single digit pg/mL.
These data demonstrate that the PHERAstar FSX is particularly advantageous for performing quantitative THUNDER™ cytokine assays.
PHERAstar^® FSX is therefore certified for use with THUNDER™ TR-FRET.