The Human Growth Hormone (HGH) Quantitative Test Kit is based on the principle of a solid phase enzyme-linked immunosorbent assay (ELISA). The concentration of HGH is directly proportional to the color intensity of the test sample. Absorbance is measured spectrophotometrically at 450 nm.
The E2 EIA Test Kit is based on the principle of competitive binding between E2 in the test specimen and E2-HRP conjugate for a constant amount of rabbit anti-Estradiol. The E2 concentration of the specimens and controls run concurrently with the standards can be calculated from the standard curve. Absorbance is measured spectrophotometrically at 450 nm.
The progesterone EIA is based on the principle of competitive binding between progesterone in the test specimen and progesterone-HRP conjugate for a constant amount of rabbit anti-progesterone. The progesterone concentration of the specimens and controls run concurrently with the standards can be calculated from the standard curve. Absorbance is measured spectrophotometrically at 450 nm.
The Testosterone EIA is based on the principle of competitive binding between Testosterone in the test specimen and Testosterone-HRP conjugate for a constant amount of rabbit anti-Testosterone. The Testosterone concentration of the specimens and controls run concurrently with the standards can be calculated from the standard curve. Absorbance is measured spectrophotometrically at 450 nm.
The High Sensitivity C-Reactive Protein (hsCRP) ELISA is based on the principle of a solid phase enzyme-linked immunosorbent assay. The concentration of CRP is directly proportional the color intensity of the test sample. Absorbance is measured spectrophotometrically at 450 nm.
This kit utilizes an Aldehyde Reactive Probe (ARP) that reacts specifically with aldehyde groups exposed in the AP sites of purified DNA samples. Following the reaction with ARP, the sites are tagged with biotin and quantified by colorimetric detection of the binding HRP-streptavidin. Each kit provides the required reagents for assessment of DNA damage in 25 purified DNA samples using a 96 well plate format.
The GSH-400 method is based on a chemical reaction which proceeds in two steps. The first step leads to the formation of substitution products (thioethers) between a patented reagent, R1 (4-chloro-1-methyl-7-trifluromethyl-quinolinium methylsulfate), and all mercaptans (RSH) which are present in the sample:
The second step is a b-elimination reaction which takes place under alkaline conditions. This reaction is mediated by reagent R2 (30% NaOH) which specifically transforms the substitution product (thioether) obtained with GSH into a chromophoric thione which has a maximal absorbance wavelength at 400 nm.
The LPO-586 Assay Kit is based on the reaction of a chromogenic reagent, N-methyl-2-phenylindole (R1), with MDA and 4-hydroxyalkenals at 45°C. One molecule of either MDA or 4-hydroxyalkenal reacts with 2 molecules of reagent R1 to yield a stable chromophore with maximal absorbance at 586 nm. For simultaneous determination of MDA and 4-hydroxyalkenals, one must use the procedure utilizing methanesulfonic acid (MSA) as the acid solvent. The procedure in which HCl is used will only detect MDA, since the 4-hydroxyalkenals do not form a chromophore with reagent R1 under those conditions.
The Lactoƒ-EIA method is an enzyme-linked immunosorbent assay (ELISA). Lactoferrin is captured by a monoclonal antibody (MAb) that is coated on wells of a sectional microplate. A second LTF-MAb labeled with biotin is added to the well and binds with the captured LTF forming a “sandwich.” A solution of streptavidin-peroxidase is then added. Streptavidin has a high affinity for biotin and once bound, its horseradish peroxidase (HRP) label is available for color development by addition of the substrate, o-phenylenediamine (OPD). This color development at 450 nm is proportional to the quantity of LTF in the sample.
The GPx-340 assay kit is an indirect measure of the activity of c-GPx. Oxidized glutathione (GSSG), produced upon reduction of an organic peroxide by c-GPx, is recycled to its reduced state by the enzyme glutathione reductase (GR). The oxidation of NADPH to NADP + is accompanied by a decrease in absorbance at 340 nm (A340) providing a spectrophotometric means for monitoring GPx enzyme activity.
The GR-340 assay is based on the oxidation of NADPH to NADP+ catalyzed by a limiting concentration of glutathione reductase. One GR activity unit is defined as the amount of enzyme catalyzing the reduction of one micromole of GSSG per minute at pH 7.6 and 25°C. Therefore, the reduction of GSSG is determined indirectly by the measurement of the consumption of NADPH, as demonstrated by a decrease in absorbance at 340 nm as a function of time.
The method is based on the formation of a chromophoric thione. The absorbance measured at 420 nm is directly proportional to the GSH concentration. There are three steps to the reaction. First, the sample is buffered and the reducing agent, tris(2-carboxyethyl)phosphine (TCEP), is added to reduce any oxidized glutathione (GSSG) to the reduced state (GSH). The chromogen, 4-chloro-1-methyl-7-trifluoromethylquinolinium methylsulfate, is added forming thioethers with all thiols present in the sample. Upon addition of base to raise the pH greater than 13, an b-elimination specific to the GSH-thioether results in the chromophoric thione.
The H2O2-560 Assay is based on the oxidation of ferrous ions (Fe2+) to ferric ions (Fe3+) by hydrogen peroxide under acidic conditions. The ferric ion binds with the indicator dye xylenol orange (3,3’-bis[N,N-di(Carboxymethyl)-aminomethyl]-o-cresolsulfone-phthalein, sodium salt) to form a stable colored complex which can be measured at 560 nm.
The Fe3+-xylenol orange complex has an extinction coefficient of 1.5 X 104 M-1•cm-1 at 560 nm when ferric ions are added in the absence of hydrogen peroxide. However, in the presence of sorbitol there is substantial chain oxidation of ferrous ion, increasing the color yield. This results in an apparent extinction coefficient of 2.67 X 105 M-1•cm-1, indicating that approximately 18 moles of Fe2+ are oxidized to Fe3+ for every mole of hydrogen peroxide present, increasing the sensitivity of the assay.
The 8-OHdG-EIA Kit is a competitive ELISA for the quantitative measurement of 8-hydroxy-1’deoxyguanosine (8-OHdG) in tissue, serum, plasma, and urine resulting from oxidative damage to DNA. 8-OHdG is produced during DNA repair and its measurement may be useful as a marker of oxidative damage due to aging, cancer, and other degenerative diseases.
The accurate measurement of GSSG levels has proved very difficult due to the low amount of GSSG in tissues and because of the absence of effective methods to prevent oxidation of GSH to GSSG during sample preparation. The GSH/GSSG-412 assay uses the thiol-scavenging reagent, 1-methyl-2-vinylpyridinium trifluoromethanesulfonate (M2VP) at a level that rapidly scavenges GSH but does not interfere with the GR assay. An enzymatic method for quantitative determination of amounts of total(reduced and oxidized or GSHt) glutathione is introduced. The method employs Ellman’s reagent (5,5'-dithiobis-2-nitrobenzoic acid or DTNB), which reacts with GSH to form a spectrophotometrically detectable product at 412 nm. GSSG can be determined by the reduction of GSSG to GSH, which is then determined by the reaction with Ellman’s reagent. In brief, the method utilizes the change in color development during the reaction, and the reaction rate is proportional to the GSH and GSSG concentrations.
The Aconitase-340 Assay is based on measurement of the concomitant formation of NADPH from NADP+. The formation of NADPH is monitored by the increase in absorbance at 340nm. Under appropriate conditions, the rate of NADPH production is proportional to aconitase activity. The concentration of aconitase is expressed in units of activity. The increase in isocitrate is equal to the consumption of NADP+, measured as the increase in absorbance at 340 nm.
The Catalase-520 assay is a two-step procedure. The rate of dismutation of hydrogen peroxide (H2O2) to water and molecular oxygen is proportional to the concentration of catalase. Therefore, the sample containing catalase is incubated in the presence of a known concentration of H2O2. After incubation for exactly one minute, the reaction is quenched with sodium azide. The amount of H2O2 remaining in the reaction mixture is then determined by the oxidative coupling reaction of 4-aminophenazone (4-aminoantipyrene, AAP) and 3,5-dichloro-2-hydroxybenzenesulfonic acid (DHBS) in the presence of H2O2 and catalyzed by horseradish peroxidase (HRP).
The resulting quinoneimine dye is measured at 520 nm (N-(4-antipyrl)-3-chloro-5-sulfonate-p-benzoquinonemonoimine).
The MDA-586 Assay Kit is based on the reaction of a chromogenic reagent, N-methyl-2-phenylindole (R1, NMPI), with MDA at 45°C. One molecule of MDA reacts with 2 molecules of NMPI to yield a stable carbocyanine dye. The concentration of MDA in an unknown is determined from the absorbance of the unknown and the standard curve.at 586 nm in the MDA-586 assay.
The Urinary 8-epi-Prostaglandin F2a Assay is a competitive enzyme-linked immunoassay (ELISA) for determining levels of 8-epi-prostaglandin-F2a in urine samples. Briefly, the samples are mixed with an enhancing reagent that essentially eliminates interferences due to non-specific binding. The 8-epi-prostaglandin-F2a in the sample or standard then competes with 8-epi-prostaglandin-F2a conjugated to horseradish peroxidase (HRP Conjugate) for binding to a polyclonal antibody specific for 8-epi-prostaglandin-F2a coated on the microplate. Following substrate addition, the intensity of the color is inversely proportional to the amount of unconjugated 8-epi-prostaglandin- F2a in the sample or standard.
The reduction potential of the sample or standard effectively converts Cu+2 to Cu+1, thus changing the ion’s absorption characteristics. This reduced form of copper will selectively form a stable 2:1 complex with the chromogenic reagent with an absorption maximum at ca. 450 nm. A known concentration of uric acid is used to create a calibration curve, with the data being expressed as mM uric acid equivalents or in μM copper reducing equivalents. A study was performed to evaluate the correlation between the concentration of principal antioxidants in human serum and the value obtained for total antioxidant power using this method. Multivariate analysis of the obtained results yields a very high significance. The results obtained with the Total Antioxidant Power Assay for a series of serum samples were also compared to the resistance to oxidation of the serum lipids in these samples. The results of this study also show that these two parameters are highly correlated. The higher the total antioxidant power, the more protected the serum lipids are from oxidation.
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