Total Antioxidant Capacity-Peroxyl Radical Assay Kit

NWLSS | Northwest Life Science Specialties, LLC.
nwk0tac01
NWK-TAC01 $995.00 each

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NWK-TAC01
The NWLSS™ TAC-Peroxyl Antioxidant Capacity kit is intended for the quantitative measurement of a sample's antioxidant status against peroxyl radical challenge. This product represents a more concise tool allowing investigators to assess a sample's antioxidant capacity against a specific, biologically relevant free radical species. The peroxyl radical is involved in lipid peroxidation and other biologically significant pathways. This product is useful for testing many types of samples including blood plasma or serum, CSF, tissue homogenates, cell lysates, seminal plasma, wine, fruit juices, brewed teas and other botanical extracts. Data generated is in Trolox equivalents similar to the popular ORAC assay.
 
Recommended for researchers wishing to assay the antioxidant capacity of various biological samples

Introduction

Reactive oxygen free radicals (ROS) have been implicated in more than 100 human diseases and in the aging process. Tissue damage caused by free radicals is also well documented in trauma, toxic shock and ischemia/reperfusion injury. ROS are generated endogenously by various pathways including aerobic respiration, inflammation and lipid peroxidation. Exogenously generated ROS pose an unprecedented challenge to organisms because of environmental deterioration, tobacco smoking, ionization radiation, UV-light exposure, organic solvents, anesthetics, pesticides and medications. Because of the ever present threat posed by exposure to ROS, organisms have developed powerful antioxidant defense systems to minimize or prevent possible deleterious effects of ROS exposure. Enzymatic systems such as superoxide dismutase, glutathione peroxidase and catalase aid in the decomposition of harmful radical species. Small molecules such as ascorbic acid, glutathione, uric acid, vitamin-E and CoQ-10 act as free radical scavengers. Macromolecules work to chelate metals and adsorb free radicals helping to further reduce possibly damaging effects. The overall antioxidant status is also related to other factors such as disease, life-style and an organism's stress load in general.
Numerous methods have been described to evaluate the total antioxidant capacity (TAC) of samples. These methods include scavenging assays that challenge a sample with superoxide anion radical, hydrogen peroxide, hypochlorous acid, hydroxyl radical, peroxyl radicals or peroxylnitrite. There are also methods using less biologically relevant systems such as those measuring a sample's capacity to reduce ferric ion and cuperic ion as well as those measuring a sample's scavenging ability toward 2,2-diphenyl-1picryhydrazyl (DPPH) radical and towards N,N-dimethyl- p-phenyleneamine (DMPD) radical. Since individual antioxidants differ in scavenging ability toward specif ROS species, it is important to note that TAC data generated using different assay platforms can vary to a significant degree. Because of this fact, it is best to describe TAC data in terms of a specific ROS challenge species.

Test Principle

The platform of the NWLSS™ TAC01 kit is an artificial system where biologically relevant peroxyl free radicals are generated by thermal decomposition of 2,2' -azobis(2-amidinopropane) (ABAP). The ABAP decomposition products are a pair of C-centered free radicals R. and a nitrogen molecule. The R. free radicals further react with oxygen molecules to form biologically relevant peroxyl radicals ROO., which are similar to those found in vivo during lipid peroxidation. These peroxyl radicals react with an indicator molecule, luminol (LH2), to generate a luminol radical (LH.) that results in emission of blue light centered at ~425 nm. When nonenzymatic antioxidants are present, luminescence is inhibited until the antioxidants are exhausted. The time of inhibition or the induction time to light production is proportional to the total concentration of antioxidants. The antioxidant concentration is determined by comparing induction time to that of a water-soluble Vitamin E (tocopherol) analog, Trolox. The principle of this assay is shown in the following scheme:

View Assay Protocol

Catalog Number: NWK-TAC01
Format: 96 tests using microtiter plate. Assay can also be performed in a single tube luminometer for a 36 test yield.
Sample Requirements: Blood plasma or serum, CSF, tissue homogenates, cell lysates, seminal plasma, wine, fruit juices, brewed teas and other botanical extracts.
Sensitivity: Lower limit of detection is 0.5 mM Trolox equivalents.
Storage and Stability: 6 months from date of manufacture if stored at 2-8C
Kit Contents: ABAP Reagent
Luminol Reagent
Trolox Standard
Assay Buffer
Sample Dilution Buffer

How does the NWK-TAC01 compare to the ORAC and FRAP assays?

The FRAP assay is different in that it relies on the reduction of metal ions. Our NWK-ARC02 assay is somewhat analogous to the FRAP (Ferric Reducing Antioxidant Power) in that both utilize a metal ion as the reductive measurement. In the case of our kit the metal ion is Cu instead of Fe. For obvious reasons we couldn't replace the Ferric with Cupric and create a new acronym similar to FRAP to describe our assay so we chose to call it an Antioxidant Reductive Capacity assay.

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