PiBlue™ Phosphate Assay Kit
Protocol SDS  Printed Protocols and SDSs are not provided with kits.

For sensitive and high-throughput phosphate determination.

Key Features
Reagent very stable. Due to our innovative formulation, no precipitation of reagent occurs. Therefore no filtration of reagent is needed prior to assays, as is often required with other commercial kits.

High sensitivity and wide detection range: detection of as little of 20 pmoles of phosphate and useful range between 0.4 μM and 50 μM phosphate.

Fast and convenient: single reagent "mix-and-measure" assay allows quantitation of free phosphate within 30 minutes.

Compatible with routine laboratory and HTS formats: assays can be performed in tubes, cuvettes or microplates, on spectrophotometers and plate readers.

Robust and amenable to HTS: Z factors of 0.7 to 0.9 are observed in 96-well plates. Can be readily automated on HTS liquid handling systems

OD620nm (malachite green)

Enzyme reactions with free phosphate releases


30 min

500 tests

Detection Limit
0.4 μM

Shelf Life
12 months

More Details
The PiBlue™ Phosphate Assay Kit is based on a proprietary formulation of the malachite green dye. The PiBlue™ reagent forms a blue colored complex with free orthophosphate. The rapid color formation from the reaction can be conveniently measured on a spectrophotometer (600 - 660 nm) or on a plate reader. The non-radioactive colorimetric assay kits have been optimized to offer superior sensitivity and prolonged shelf life. The assay is simple and fast, involving a single addition step for phosphate determination. Assays can be performed in tubes, cuvettes or multi-well plates. The assays can be conveniently executed in 96-well plates for high-throughput screening of enzyme inhibitors.

1. What is the difference between your 3 phosphate assays: DIPI-500, POMG-25H, POPB-500?

All three products are based on the malachite green dye. While the PiBlue (POPB-500) and QuantiChrom (DIPI-500) Phosphate Assay Reagents are identical, they differ in the standards. The PiBlue is designed for customers who want prepare a full calibration curve for their assays, and the latter has one set of a blank control and 30 µM phosphate standard. The reagent is more diluted than the reagent used in the Malachite Green Phosphate Assay Kit (POMG-25H). As a result, one adds 100 µL reagent (POPB-500 or DIPI-500) to 50 µL sample, whereas for the Malachite Green based assay, one adds 20 µL Reagent to 80 µL sample.

In practice, the DIPI-500 is the most convenient kit to use, because there is no need to prepare a calibration curve there is no need for extra mixing. Because adding a larger volume (100 μL) to a small volume (50 μL) effects efficient mixing in well, no extra mixing is needed. The POPB-500 requires preparation of calibration curve. With the POMG-25H Kit, one needs to mix the reagent with the sample, because adding 20 µL to 80 µL does not mix efficiently by itself. Also a preparation of a standard curve is necessary.

For more detailed product information and questions, please feel free to Contact Us. Or for more general information regarding our assays, please refer to our General Questions
1. Lau KH et al (2006). An osteoclastic protein-tyrosine phosphatase is a potential positive regulator of the c-Src protein-tyrosine kinase activity: a mediator of osteoclast activity. J Cell Biochem. 97(5):940-55. Assay: Phosphate in Mouse cytosolic fraction (Pubmed).

2. Aldana-Masangkay GI et al (2011). Tubacin suppresses proliferation and induces apoptosis of acute lymphoblastic leukemia cells. Leuk Lymphoma. 52(8):1544-55. Assay: Phosphate in Rabbit phosphatases (Pubmed).

3. Liu Q et al (2011). Competition between foliar Neotyphodium lolii endophytes and mycorrhizal Glomus spp. fungi in Lolium perenne depends on resource supply and host carbohydrate content. Functional Ecology 25:910–920. Assay: Phosphate in Mouse intracellular phosphate (Pubmed).

4. Chen X et al (2007). The isolation and structure of membrane lipid rafts from rat brain. Biochimie 89(2):192-6. Assay: Phosphate in mouse/human cellular extract (Pubmed).

5. Hough TA et al (2007). Novel mouse model of autosomal semidominant adult ypophosphatasia has a splice site mutation in the tissue nonspecific alkaline phosphatase gene Akp2. J Bone Miner Res. 22(9):1397-407. Assay: Phosphate in Frog Na+/K+-ATPase extract (Pubmed).

6. Kang HY et al (2008). Altered TNSALP expression and phosphate regulation contribute to reduced mineralization in mice lacking androgen receptor. Mol Cell Biol.28(24):7354-67. Assay: Phosphate in Rat ATPase extract (Pubmed).

7. Cramp RL et al (2009). Ups and downs of intestinal function with prolonged fasting during aestivation in the burrowing frog, Cyclorana alboguttata. J Exp Biol. 212(22):3656-63. Assay: Phosphate in Mouse lipid Pi (Pubmed).

8. Demidenko AA et al (2011). Effects of viscogens on RNA transcription inside reovirus particles. J Biol Chem. 2011 Jun 30. Assay: Phosphate in Rat lipid (Pubmed).

9. Polewski MD et al (2010). Inorganic pyrophosphatase induces type I collagen in osteoblasts. Bone 46(1):81-90. Assay: Phosphate in Human blood, semen (Pubmed).

To find more recent publications, please click here. To find more recent publications, please click here.
If you or your labs do not have the equipment or scientists necessary to run this assay, BioAssay Systems can perform the service for you.

- Fast turnaround
- Quality data
- Low cost

Please inquire via email or call 1-510-782-9988 x 2.
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PiBlue™ Phosphate Assay Kit
Catalog No: POPB-500
Price: $189    Qty:
For orders of 10 or more kits, please call 1-510-7829988x1 or email us for best pricing and/or bulk order.

Shipping: RT
Shipment: Fedex Service
Delivery: 1-2 days (US), 3-6 days (Intl) Storage: 4°C

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