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PiColorLock™ Phosphate Detection System

PiColorLock™ phosphate detection reagent

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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.

PiColorLock™ - phosphate detection reagent features and benefits

Features Benefits
Colorimetric assay Simple, scalable and non-radioactive
Compatible with almost any assay buffer Flexibility over a wide range of existing assays
Stable reagent formulation Long shelf life
Unique accelerator Speeds up color development
Unique stabilizer Suppresses non enzymatic backgrounds with acid-labile substrates (ATP or GTP)
PiColorLock™-Pi complex is very stable                       No precipitation, results can be measured over several hours – ideal for HTS
Wide linear range No inhibition of color development by high concentrations of protein

The unique PiColorLockTM Stabilizer prevents background drift with time

PiColorLock Data Image

The PiColorLockTM reagent is often used with unstable substrates (e.g. ATP, GTP) that give rise to non-enzymatic background drift with time. The unique stabilizer, provided in the PiColorLockTM phosphate detection system, blocks this non-enzymatic breakdown generating a stable low background. The stable signal can therefore be read up to several hours after the reaction has ended.


For more information on PiColorLock™ take a look at our Phosphate Detection Guide.

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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).

assay background web

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.

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.

Which reagents does the PiColorLock™ Phosphate Detection System contain?

Reagent Product Code: 303-0030 Product Code: 303-0125
PiColorLock 30ml 125ml
Stabilizer 27.5ml 55ml
Accelerator 1ml 2ml
Phosphate Standard 5ml 11ml
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PiColorLock - 625/1560 assays (303-0030)

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PiColorLock - 2500/6250 assays (303-0125)

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Our shipping times are as follows:
48 hours to the US. 24 to 48 hours to Europe.

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