• Detection method- Absorbance (650 nm)
• Sample type- Cell and Tissue lysates, culture media, urine, plasma and serum, as well as many other biological fluids, algal blooms and water from run-off areas of high fertilizer use
• Species reactivity- Mammalian
• Applications- Phosphate concentrations between 1 µM and 1 mM, with a lower limit of detection of approximately 0.1 nmol, can be directly determined.
Features & Benefits:
• Simple procedure; takes ~ 30 minutes
• Fast and convenient
• The assay is sensitive, stable and high-throughput adaptable
• Phosphate Reagent
• Phosphate Standard (10 mM)
Phosphate is one of the most important of the inorganic ions in biological systems. It functions in a variety of roles. One of the most important roles is as a molecular switch, turning enzyme activity on and off through the mediation of the various protein kinases and phosphatases in biological systems. Phosphate is also of great importance in mineralization processes and is a primary stimulus of algal blooms frequently found in bodies of fresh water, due to run-off from areas of high fertilizer use. The newly designed Phosphate Colorimetric Assay Kit provides an easy, quick and sensitive means of assessing phosphate over a wide range of concentrations. The assay utilizes a proprietary formulation of malachite green and ammonium molybdate which forms a chromogenic complex with phosphate ion giving an intense absorption band around 650 nm. Phosphate concentrations between 1 µM and 1 mM, with a lower limit of detection of approximately 0.1 nmol, can be directly determined. The Phosphate Colorimetric Assay Kit provides 500 assays using microtiter plates or 100 assays using 1 ml cuvettes.
USAGE: For Research Use Only! Not For Use in Humans.
We want to perform this assay and are worried about the influence of the precipitates on the OD reading. How should we proceed?
Any precipitate in the wells will interfere with the reading and that has to be avoided. As long as you are not pipetting in the precipitate and taking it into the analysis wells, it should be fine. The clear reagent can be used directly for the assay.
We would like to use salt water based media with approx. 17,000 ppm of chloride, it will not inhibit the reaction or have any other adverse effects or interference the data, right?
Our development scientist has confirmed that the chloride in the samples will have no interference in the assay.
My purpose is to check the amount of phosphate uptake in the cells before and after certain treatment. Can I use this kit to for this purpose? What protocol do you recommend? Any special treatment for lysing the cell?
Yes, this kit can be used for your purpose. Use any generic lysis buffer for making the extract. Use a Dounce homogenizer / sonicator for homogenization. Cell and tissue extracts/lysates also contain phosphates from the breakdown of nucleic acids, lipids, etc. Extraneous free phosphate should be removed from samples to be analyzed. Two commonly used methods of removing phosphate from samples are desalting columns and immunoprecipitation capture of thePhosphate-generating enzymes being studied.
I am looking for a commercial kit for quantifying the amount of phospholipid in some liposome samples. The most frequent used method is to determine the content of phosphor in the sample, and thereby determine the amount of phospholipid. I was considering if I could use your Phosphate Colorimetric Assay kit for this application. Would you be able to measure th phosphor amount directly on the intact phospholipids, or does the liposome suspension have to be pretreated in order liberate the phosphate as a free ion?
The phosphate assays colorimetric and fluorometric measure inorganic phosphate, so you would need to release the phosphate enzymatically from your liposomes.
Regarding your kit 410-500, I would like to know what is your standard and what type of phosphate the kit reads: PO4(3-), H2PO4 (-), HPO4(2-), …? Or all of them, even when you have a mixture of those salts?
The kit will detect all the three types of PO4 in the reaction.
Can this kit be used with samples like bacteria, plants, drosophila, yeast etc?
We have optimized the kit with mammalian samples. However, theoretically these kits should work with samples from multiple species/sources. Since the optimal conditions depend on the sample type, the protocol has to be be adapted to fit the samples for efficient results. Please refer to this kits citations to see what kind of samples have been used with this kit other than mammalian samples.
Can we use frozen samples with this assay?
Fresh samples are always preferred over frozen samples. However, frozen samples can also be used, provided, they were frozen right after isolation, were not freeze thawed multiple time (for which we recommend aliquoting the samples before freezing) and have been frozen for relatively short periods.
Can we use a different wavelength than recommended for the final analysis?
It is always recommended to use the exact recommended wavelength for the most efficient results. However, most plate readers have flexibility in their band width of detection in increments of +/- 10 nm. Depending on this flexibility range, you can deviate from the recommended wavelengths within limits.
What is the exact volume of sample required for this assay?
There is no specific volume we can recommend for the amount any sample to be used since it is completely sample concentration and quality based. You have to do a pilot expt with multiple sample volumes to determine the optimal volume which gives a reading within the linear range of the standard curve. Please refer to the citations for this product to see what other clients have used with similar sample types.
Do you have trial sizes of this kit?
Unfortunately, we do not have trial sizes of this kit available. However, if you are based in the US or Canada, we will give you a 10% off list price introductory discount on its purchase price. If you are based out of this area please contact yopur regional BioVision distributor.
What is the shelf life of this kit?
This kit is good for 12 months from the date of shipment.
Can we purchase individual components of this kit?
Yes, you can purchase any of the kits components without the whole kit. Please refer to the component Cat #s mentioned on the datasheet for ordering.
Should I make a standard curve for every expt I do, or is one curve/kit enough?
Yes, I would strongly recommend you to do the standards every time you do the expt. There is always a chance that something was done differently that day and we do not want any conditions to differ between standards and samples.