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Aldehyde Dehydrogenase Activity Colorimetric Assay Kit

based on 1 citations in multiple journalsAldehyde Dehydrogenase Activity Colorimetric Assay Kit14.1 4
Catalog #: K731

In stock

$455.00

Product Details

Cat # +Size K731-100
Size 100 assays
Detection Method Absorbance (450 nm)
Species Reactivity Mammalian
Applications The assay can detect < 0.5 mU of ALDH activity (based on our unit definition) in a variety of samples.
Features & Benefits • Simple procedure
• Fast and convenient
• Sensitive assay for measuring ALDH activity in various biological samples and the kit is also suitable for high throughput analyses.
Kit Components • ALDH Assay Buffer
• Acetaldehyde
• ALDH Substrate Mix (Lyophilized)
• ALDH Positive Control (Lyophilized)
• NADH Standard (0.5 µmol, Lyophilized)
Storage Conditions -20°C
Shipping Conditions Gel Pack
USAGE For Research Use Only! Not For Use in Humans.

Details

The NAD-dependent Aldehyde Dehydrogenase (ALDH) plays a vital role in cellular detoxification. It oxidizes various aldehydes and generates the corresponding carboxyolic acid. ALDH have been found in every cellular compar™ent. Based on its structure and function, ALDH comprises 3 major classes in mammals: Class 1 and Class 3 (the tumor form) are located in the cytosol and include both constitutive and induced forms; Class 2 is located in the mitochondria and only exists as the constitutive form. In humans, the ALDH superfamily consists of 19 genes. The mutation of ALDH genes (loss of function) causes human diseases such as Type II hyperprolinemia, pyridoxine-dependent seizure and hyperammonemia. Recent studies show that increased ALDH activity leads to several types of malignancies, serves as a cancer stem cell marker and correlates with poor prognosis. Therefore the early detection of ALDH activity levels can be prognostic and guide the therapeutic strategies. The BioVision Aldehyde Dehydrogenase (ALDH) Activity Assay Kit is a simple, fast and reliable method to quantify the ALDH enzymatic activity. In this assay, acetaldehyde is oxidized by ALDH generating NADH which then reduces a colorless probe to a colored product with strong absorbance at 450 nm. The assay can detect < 0.5 mU of ALDH activity (based on our unit definition) in a variety of samples.


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Can you describe the positive control used in the kit?
The positive control is an ALDH isolated from Baker’s yeast.
Is ALDH activity kit compatible with cell lysates prepared using RIPA buffer or other similar lysis buffers? If not, does your Assay Buffer serve as a lysis buffer or does the sample need to be mechanically homogenized? Also, can the lysates made in your Assay Buffer be used in other protein assays, such as Western blot or ELISA?
We would recommend using our assay buffer for the lysis, as RIPA may not contain all the components required for the reaction to occur. The samples should be homogenized in ice cold assay buffer for 10 minutes on ice (page 8 of the online protocol). Lysates prepared in the assay buffer can also be used in Western blot and ELISA.
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 kit's 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 kit's 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, 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.
How to remove NADPH and NADH from the samples?
The 10kda spin filter can be used to remove NADPH or NADH from the samples, but collect the upper retained fraction and use it for the assay. It could be very viscous, so further dilution is necessary. For calculation purpose, they need to remember the original volume of the sample loaded and factor that in while doing data analysis.
What is the final volume after adding the 50 uL reaction mix? 100 uL or 50 uL?
There is 50ul from sample + assay buffer and then 50ul reaction mix making the total volume =100ul.
What is the source of the positive control?
The positive control is from yeast.
whether the 20% glycerol means volume ratio? So I should add 220 ul Assay Buffer and 55 ul glycerol in the vial to prepare the ALDH Positive Control ,right?
20% glycerol is by volume. You need to add 55ul glycerol and 220ul buffer.
How to detect ALDH activity located in the mitochondria?
This kit detects ALDH based on its ability to oxidize acetaldehyde. To detect ALDH2 specifically, isolate mitochondria from the cells (please check out BioVision’s mitochondria isolation kit. http://www.biovision.com/mitochondria-isolation-kit-for-tissue-cultured-cells-7003.html) and then do the assay as per the kit protocol.
Choi NE et al., (2017) Effects of Cudrania Tricuspidata Root Extract (CTE) on Ethanol-Induced Hangover via Modulating Alcohol Metabolizing Enzyme Activities and Blood Gas Levels in Rats, Journal of the Korea Academia-Industrial cooperation Society, 2017, 18(2):218-225
For more citations of this product click here
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