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Evaluation of PARP inhibitors: performed on BMG LABTECH's FLUOstar Omega

Rebecca Foster, Kyla Grimshaw Hypoxium Ltd. 04/2008

PARP enzymes transfer ADP-ribose onto proteins and extend them to poly-ADP-ribose (PAR) branches. This way, sites of single-strand DNA breaks are marked at their surrounding histones and repair of the lesions is accelerated. The enzyme is blocked in therapy of cancers restricted in further DNA repair pathways.

 

A PARP inhibitor was tested regarding its inhibitory action in cell lysate and regarding its cytotoxic effect in LoVo adenocarcinoma cells. All readouts were done on a FLUOstar® Omega multi-mode plate reader.

 

Cell lysates were adjusted after protein quantification with the colorimetric BCA assay. The IC50 of the investigated PARP-inhibitor was 14 nM, as determined with the chemiluminescent PARP-activity assay of Trevigen®. The inhibitor has further proven to act synergistically in cell killing with the DNA-damaging chemotherapeutic Temozolomide. The cell viability was assessed with the fluorescent AlamarBlue® assay.

 

The ability of the FLUOstar Omega to measure absorbance, luminescence and fluorescence, facilitates simple, rapid measurement of all aspects of our evaluation, using a single machine.

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