PiColorLock Phosphate Detection System
PiColorLock™ phosphate detection reagent
PiColorLock™ phosphate detection reagent changes colour in the presence of inorganic phosphate (Pi). This can be exploited to measure any enzyme that generates Pi including ATPases, GTPases, phosphatases, heat-shock proteins and DNA unwinding proteins.
The unique formulation of PiColorLock affords enhanced assay linearity, dynamic range and colour stability; it is also possible with PiColorLock to work with unstable substrates (e.g. ATP, GTP) that give high non-enzymatic background signals with other acidic dye-based detection reagents.
Key Features of PiColorLock™ - phosphate detection reagent include:
- 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.
For more information on PiColorLock take a look at our Phosphate Detection Guide.
Figure 1. Competitor assays are beset with several problems including reagent precipitation. The high stability of PiColorLock™ ensures high stability of the colored dye-phosphate complexes (green color).
Figure 2. ATP has been incubated in three detection reagents. A steadily rising background signal is seen with competitor reagents, whereas PiColorLock™ gives baseline readings.
Figure 3. PiColorLock has also been designed to have a large linear range, thus reducing the need for sample dilution. For the purposes of comparison, data for PiColorLock ALS reagent (no longer available) is also shown, as it provides similar absorbance values to competitor products, at least at low levels of inorganic phosphate. Competitors’ products are linear over a much narrower range of concentrations.
For more information, please watch our webinar on drug screening-assays for phosphate-generating enzymes below:
I have a high background in my ATPase assay and I definitely do not have free phosphate in my sample
This is almost always due to inadequate mixing of the special stabiliser with the sample and detection reagent. Make sure the stabiliser is pipetted up and down several times to ensure thorough mixing.
I would like to measure the conversion of pyrophosphate to phosphate. Can I use the PiColorLock Phosphate Detection System for this purpose?
Yes, only the phosphate will give a signal; pyrophosphate will not.
I have 5% DMSO in my assay. Can I use PiColorLock?
Yes, the reagent is designed for drug screening work and other situations that require DMSO.
I have phosphate in my enzyme. What can I do?
You can dialyse or desalt the enzyme into a phosphate-free buffer. Alternatively, you can use a special resin (PiBind) to remove the phosphate.
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