High-throughput screening assays utilizing metal-chelate capture
AssigneeMitotix, Inc.
InventorsSerene Josiah, Michael Boisclair
Life of this patent
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Abstract
A high throughput enzyme screen has been developed which relies on metal chelate interaction for capture of the product of the enzymatic reaction. In the present assay system, a detectable moiety is attached to a substrate having a chelating capturable moiety, which can be captured by an immobilized metal. Detection is effected due to the presence of a detectable label on the reaction product immobilized on the solid phase. Only signal associated with tagged protein bound to the solid phase is detected. The present assay can reliably measure enzyme activity, and has high reproducibility, which benefits high throughput screening.
Claims19
An assay method for determining the effect of a substance on the activity of an enzyme, comprising the steps of:
- a. combining a test compound with an enzyme capable of catalyzing the transfer of a detectable moiety to a substrate, a donor substance labeled with the detectable label, and a substrate specific for the enzyme comprising a metal-chelating moiety, and incubating the mixture under conditions sufficient to promote the enzymatic reaction;
- b. contacting the combination formed in (a) with a metal-derivatized solid phase under conditions sufficient to promote reaction of the immobilized metal and the chelating moiety;
- and
- c. detecting the signal generated.
An assay method for determining the effect of a substance on the activity of a prenyl transferase enzyme, comprising the steps of:
- a. combining a test compound with a prenyl transferase enzyme, a radioactively-labeled donor substance, and a substrate specific for the prenyl transferase enzyme comprising a metal-chelating moiety, and incubating the mixture under conditions sufficient to promote the enzymatic reaction;
- b. contacting the combination formed in (a) with a metal-derivatized solid phase under conditions sufficient to promote reaction of the immobilized metal and the chelating moiety;
- and
- c. detecting the signal generated.
Dependencies are inferred from the claim text.