UREA ASSAY KIT
• Detection method- Absorbance (570 nm)
• Sample type- Cell and Tissue lysates, culture media, urine, plasma and serum, as well as many other biological fluids
• Species reactivity- Mammalian
• Applications- The kit can detect as low as 0.5 nmol per well or 10 µM of Urea
Features & Benefits:
• Simple procedure; takes ~ 1 hour
• Fast and convenient
• The assay is sensitive, stable and high-throughput adaptable.
• Urea Assay Buffer
• OxiRed Probe in DMSO
• Enzyme Mix (lyophilized)
• Converting Enzyme
• Urea Standard (100mM)
Urea is a waste product which produced in the liver, dissolved in blood (in a concentration of 2.5 - 7.5 mM), and secreted by the kidneys. Urea also plays a very important role in protein catabolism, removal of toxic ammonia from the body, and the countercurrent system which allows for reabsorption of water and critical ions in the nephrons. Urea determination is very useful for the medical clinician to assess kidney and other organs function of patients. BioVision’s Urea Assay Kit provides a rapid, simple, sensitive, and reliable for measurement of Urea level in a variety of samples such as serum, plasma, and urine, etc. In the assay, Urea reacts as substrate with compounds in the presence of enzymes to form a product that reacts with the OxiRed probe to generate color (λmax=570nm). The optical density of produced color has a direct relationship with Urea concentration in the solution. The kit can detect as low as 0.5 nmol per well or 10 µM of Urea. The assay is also suitable for high throughput studies.
USAGE: For Research Use Only! Not For Use in Humans.
As my samples (mouse urine) need 500-fold dilution there is too little assay buffer in the kit. I dilute 1uL sample in 999uL assay buffer and from this dilution I use 2 uL for each well. Hereafter 48uL of assay buffer is added according to the protocol. Would it be possible to use dH20 instead of assay buffer for the initial 1:1000 dilution?
Yes, for this one particular case, since you are using so little of the diluted sample and making us the volume with our assay buffer, we think your initial dilution with water should be ok. But again, that is going to be restricted for this one particular case. If you were to use any more of your initially diluted samples and then make up the volume with any lesser amount of the assay buffer we would strongly recommend you to just purchase extras of the assay buffer itself with the Cat # K375-100-1.
Could I read the assay at 540 rather than 570? Does hemolysis affect the reading? What volumes would be needed of mouse serum samples?
If the detection band width on the plate reader is ± 30, then a wavelength of ± 30 of 570 can be used, which is 540-600 nm. Otherwise the efficiency of detection of this assay will decrease.
We do not have data showing that hemolysis would affect the reading, but we would think that a cleaner serum sample would give better results.
There is no exact amount of mouse serum which will be right for this assay. For unknown samples, we suggest testing several doses of your sample to ensure the readings are within the standard curve range. Make a dose curve initially to determine the optimal starting sample volume.
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 in the unopened form when stored at the appropriate temperature and appropriate conditions. After opening and reconstitution, some of the components in this kit are good for 2 months at -20°C. Please refer to the datasheet for storage information and shelf life of each of the components.
Why are my standard curve values lower than those shown on the datasheet?
There are multiple factors which influence the signals like the incubation times, room temperature, handling etc. In general, to increase the value of the standards, you can increase the incubation time. As long as the standard curve is linear, it should be fine to use, since all of your samples will also be measured under the same conditions on this curve.
How do I normalize my samples against protein concentration?
You can use a protein quantitation assay on the supernatants you get from cell/tissue lysates or with any other liquid sample in the assay buffer.
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.
Can we use an alternate buffer for sample preparation (cell lysis, sample dilutions etc)?
Our assay buffers are optimized for the reactions they are designed for. They not only contain some detergents for efficient lysis of your cells/tissue, but also contain some proprietary components required for the further reactions. Therefore, we highly recommend using the buffers provided in the kit for the best results.
Should I make a standard curve for every expt I do, or is one curve/kit enough?
Yes, It is recommended 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.