Colorimetrics-Fluorometrics
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Ready-to-use colorimetric substrates for members of caspase family proteases. All substrates were provided in liquid ready-to-use form.
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Caspase family of proteases are the central mediators of apoptosis in mammalian cells. The Caspase-12 Fluorometric Assay Kit provides a simple and convenient means for assaying the activity of caspases that recognize the sequence ATAD. The assay is based on detection of cleavage of substrate ATAD-AFC (AFC: 7-amino-4-trifluoromethyl coumarin). ATAD-AFC emits blue light (?max = 400 nm); upon cleavage of the substrate by caspase-12 or related caspases, free AFC emits a yellow-green fluorescence (?max = 505 nm), which can be quantified using a fluorometer or a fluorecence microtiter plate reader.
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Caspase family of proteases are the central mediators of apoptosis in mammalian cells. The Caspase-12 Fluorometric Assay Kit provides a simple and convenient means for assaying the activity of caspases that recognize the sequence ATAD. The assay is based on detection of cleavage of substrate ATAD-AFC (AFC: 7-amino-4-trifluoromethyl coumarin). ATAD-AFC emits blue light (?max = 400 nm); upon cleavage of the substrate by caspase-12 or related caspases, free AFC emits a yellow-green fluorescence (?max = 505 nm), which can be quantified using a fluorometer or a fluorecence microtiter plate reader.
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Glutathione S-transferase (GST) is a family of enzymes that play an important role in detoxification of xenobiotics. GST catalyzes the formation of the thiol group of glutathione to electrophilic xenobiotics. It utilizes glutathione to scavenge potentially toxic compounds including those produced as a result of oxidative stress and is part of the defense mechanism against the mutagenic, carcinogenic and toxic effects of such compounds. The GST Colorimetric Activity Assay Kit is based upon the GST-catalyzed reaction between GSH and the GST substrate, CDNB (1-chloro-2,4-dinitrobenzene, which has the broadest range of isozyme detectability (e.g., alpha-, mu-, pi-, and other GST isoforms). Under certain conditions, the interaction between glutathione and CDNB is totally dependent on the presence of active GST. The GST-catalyzed formation of GS-DNB produces a dinitrophenyl thioether which can be detected by spectrophotometer at 340 nm. One unit of GST activity is defined as the amount of enzyme producing 1 ?mol of GS-DNB conjugate/min under the conditions of the assay. The kit can detect GST activity in crude cell lysate or purified protein fraction, and also quantitate GST-tagged fusion protein. Detect limit: Active GST < 4 mU.
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Inhibition of histone deacetylase (HDAC) has been implicated to modulate transcription and induce apoptosis or differentiation in cancer cells. However, screening compounds for HDAC inhibition has been difficult due to the lack of convenient tools for analyzing HDAC activity. The Fluorometric HDAC Activity Assay Kit provides a fast and fluorescence-based method that eliminates radioactivity, extractions, or chromatography, as used in traditional assays. The new procedure requires only two easy steps, both performed on the same microtiter plate. First, the HDAC substrate, which comprises an acetylated lysine side chain, is incubated with a sample containing HDAC activity (e.g., HeLa nuclear extract). Deacetylation of the substrate sensitizes the substrate, so that further treatment with the Lysine Developer produces a fluorophore. The fluorophore can be easily analyzed using a fluorescence plate reader or a fluorometer. The assay is well suited for high throughput screening applications. HDAC inhibitors and antibodies are also available separately.
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Inhibition of histone deacetylases (HDACs) has been implicated to modulate transcription and to induce apoptosis or differentiation in cancer cells. However, screening HDAC inhibitory compounds has proven to be difficult over the past due to the lack of convenient tools for analyzing HDAC activity. The new Colorimetric HDAC Activity Assay Kit provides a fast and convenient colorimetric method that eliminates radioactivity, extractions, or chromatography, as used in the traditional assays. The new method requires only two easy steps, both performed on the same microtiter plate. First, the HDAC colorimetric substrate, which comprises an acetylated lysine side chain, is incubated with a sample containing HDAC activity (e.g., HeLa nuclear extract or your own samples). Deacetylation of the substrate sensitizes the substrate, so that, in the second step, treatment with the Lysine Developer produces a chromophore. The chromophore can be easily analyzed using an ELISA plate reader or spectrophotometer. The assay is well suited for high throughput screening applications. HDAC inhibitors and antibodies are also available separately.
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ATP is the primary energy currency of living systems. Virtually all energy requiring processes utilize the chemical energy stored in the phosphate bond of ATP. ATP is formed exclusively in the mitochondria and a variety of genetic diseases can affect ATP formation in the mitochondria. There are a number of commercially available ATP assays which detects femtomoles or less of ATP by measuring luminescence (BioVision Kit 254-200, for example) but these kits require specialized luminescence instrumentation and utilize luciferase which can be difficult to maintain in active form. BioVision newly developed ATP Colorimetric and Fluorometric Assay kit is designed to be a robust, simple method which utilizes the phosphorylation of glycerol to generate a product that is easily quantified by colorimetric (?max = 570 nm) or fluorometric (Ex/Em = 535/587 nm) methods. The assay can detect as low as 50 picomol (1 ?M) of ATP in various samples. The kit provides sufficient reagents for 100 assays.
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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 ?M 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.
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