High Throughput Colorimetric GTPase Assays

Key Features of PiColorLock™ - phosphate detection reagent:

• Colorimetric Assay – competitor assays are radioactive.
• Special additive ensures low backgrounds with acid-labile substrates.
• Unparalleled stability of phosphate-dye complexes.
• Reagent is compatible with almost any assay buffer.
• No inhibition of color development by high concentrations of protein.
• Stable reagent formulation - long shelf life.

Introduction

Colorimetric assays for GTPases are invariably based on the formation of colored complexes between an inorganic phosphate and a dye molecule under acidic conditions.

Competitor assays are beset with several problems including reagent precipitation.  At the heart of Innova Biosciences assay kit is PiColorLock™ Gold a non-radioactive, superior phosphate detection reagent which ensures high stability of the colored dye-phosphate complexes (green color- see below).

PicolorLock™ Gold has also been designed to:

• Prevent background signals arising from non-enzymatic (i.e. acid) hydrolysis of the substrate. The figure below shows ATP incubated in three detection reagents. A steadily rising background signal is seen with competitor reagents, whereas PiColorLock™ gives baseline readings.

• Have a large linear range, thus reducing the need for sample dilution.

Together with specially purified substrates and proprietary stabilizers Innova Biosciences kits offer the lowest possible assay background and outstanding assay performance. Uniquely, Innova Biosciences GTPase assay kits contain highly purified GTP, which have had any free inorganic phosphate removed – this means the assays have the lowest possible background signal.

PiColorLock™ Gold can also be purchased for developing simple assays for any phosphate-generating enzyme.

Related Reagents

PiBind™ resin provides a quick and easy way to remove contaminating Pi from buffers and from samples derived from cells and tissues.