News

Excellent performance of THUNDER™ phospho-protein and cytokine TR-FRET assays on BMG LABTECH’s CLARIOstar® Plus

2022-03-16

In the framework of our ongoing strategic collaboration with BMG LABTECH, we have evaluated the performance of a panel of three THUNDER™ TR-FRET phospho-proteins assays and five human cytokine assays on the flexible and powerful CLARIOstar^® Plus multi-mode microplate reader. The results, summarized in the Application Note 361 and in a tutorial showed an excellent assay performance of our assays on this very sensitive instrument.

Fig. 2: THUNDER™ Phospho-ERK1/2 (T202/Y204) assay (4 hours incubation).

Fig. 3: THUNDER™ Phospho-p38αβγ (T180/Y182) assay (overnight incubation).

Fig. 4: THUNDER™ Phospho-STAT3 (Y705) assay (overnight incubation).

Table 1: Performance of THUNDER™ human cytokine assays on the CLARIOstar Plus (xenon flash lamp) and the competing HTS reader (TRF laser).

Analyte	Assay parameter	CLARIOstar Plus	Competitor
IFNγ	LOD (pg/mL)	4	10
	LLOQ (pg/mL)	22	70
	S/B	11	9
IL-1β	LOD (pg/mL)	7	36
	LLOQ (pg/mL)	42	164
	S/B	23	18
IL-12	LOD (pg/mL)	4	38
	LLOQ (pg/mL)	25	179
	S/B	19	15
TNFα	LOD (pg/mL)	3	41
	LLOQ (pg/mL)	28	211
	S/B	58	42
CCL2	LOD (pg/mL)	9	44
	LLOQ (pg/mL)	40	172
	S/B	11	9

Highlights

Human IFN-gamma, IL1-beta, IL-12, CCL2, and TNF-alpha assay kits were tested in a 384-well format using the standardized protocols.
Phospho-ERK1/2 (T202/Y204), Phospho-p38αβγ1/2 (T180/Y182), and Phospho-STAT3 (Y705) assay kits were tested in a 384-well format using the 2-plate (transfer) protocol.
Assays were read on the CLARIOstar Plus equipped with a high energy xenon flash lamp excitation source.
For comparison, plates were also read on a competing HTS microplate reader equipped for TRF laser excitation.
For cytokine assays, 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 CLARIOstar Plus exhibited broader dynamic ranges, higher sensitivity and S/B ratios compared to the competitor and our internal data, decreasing the LOD of cytokines to single digit pg/mL.
The combination of simple, rapid, and robust yet affordable THUNDER™ TR-FRET assays with the highly sensitive CLARIOstar Plus provides a powerful approach for accelerating the workflow of drug discovery and life science research.
CLARIOstar^® Plus is therefore certified for use with THUNDER™ TR-FRET.

THUNDER certification