Simultaneous dual emission detection of luciferase reporter assays

Megan Dobbs (1), Douglas Hughes (1), Janaki Narahari (1), Jae Choi (1), Georgyi Los (1), Brian Webb (1), Eric Matthews (2), Carl Peters (2) (1) Thermo Fisher Scientific, (2) BMG LABTECH 03/2013

Promoter activity is often investigated using luciferases whose genes are cloned into a reporter expression vector. Conventional assays detect the light produced by a luciferase controlled by an experimental promoter and its substrate. Upon addition of a quencher of this first reaction, the signal is related to the light produced by a second control luciferase and its substrate. Here it is shown how spectrally resolved luciferases can be measured simultaneously on a POLARstar® Omega without the need for quencher-addition or separate reads.

Dilutions of HEK293 cell extracts expressing either Cypridina or Gaussia luciferase were measured in presence of red firefly luciferase mimicking the product of a control reporter. Simultaneous measurement of Gaussia and Red Firefly luminescence detected changes in Gaussia luciferase concentration while not impacting on the signal of Red Firefly. Comparable results were obtained using Cypridina instead of Gaussia luciferase.

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