Substrates

24 respective 100 ready-to-use para-Nitrophenylphosphate (pNPP) tablets with 5mg p-Nitrophenylphosphate. Each tablet dissolved in 5mL deionized water yields a solution containing 1.0mg/mL pNPP, 0.2M Tris buffer. Genaxxon's pNPP substrate is specifically developed for immunoassay procedures and it is ideal for phosphate-based ELISA methods. The substrate is also used as a chromogenic non-specific substrate in alkaline and acid phosphatase assays. Its soluble end-product is yellow and can be spectrophotometrically read at 405 to 410 nm. The reaction may be stopped with 3M NaOH. The pNPP substrate is supplied as pre-weighed tablets in bottles or in convenient blister packs, each tablet containing 5mg or 20mg of substrate. How to use:Deposit one tablet of the pNPP substrate in a laboratory flask or beaker placed on a magnetic stirrer. Add deionized water in the appropriate amount to reach the required concentration for the assay. Stir until full dissolution and the substrate solution is ready to use. One tablet dissolved in 5mL of substrate buffer results in a 1mg/mL solution.

24 respective 100 ready-to-use p-Nitrophenylphosphate (pNPP) tablets with 20mg p-Nitrophenylphosphate. Each tablet dissolved in 20mL deionized water yields a solution containing 1.0mg/mL pNPP, 0.2M Tris buffer. Genaxxon's pNPP substrate is specifically developed for immunoassay procedures and it is ideal for phosphate-based ELISA methods. The substrate is also used as a chromogenic non-specific substrate in alkaline and acid phosphatase assays. Its soluble end-product is yellow and can be spectrophotometrically read at 405 to 410 nm. The reaction may be stopped with 3M NaOH. The pNPP substrate is supplied as pre-weighed tablets in bottles or in convenient blister packs, each tablet containing 5mg or 20mg of substrate. How to use:Deposit one tablet of the pNPP substrate in a laboratory flask or beaker placed on a magnetic stirrer. Add deionized water in the appropriate amount to reach the required concentration for the assay. Stir until full dissolution and the substrate solution is ready to use. One tablet dissolved in 20mL of substrate buffer results in a 1mg/mL solution.

Medicago’s pNPP substrate is specifically developed for immunoassay procedures and it is ideal for phosphate-based ELISA methods. The substrate is also used as a chromogenic non-specific substrate in alkaline and acid phosphatase assays. Its soluble end-product is yellow and can be spectrophotometrically read at 405 to 410 nm. The reaction may be stopped with 3 M NaOH. The pNPP substrate is supplied as pre-weighed tablets in bottles or in convenient blister packs, each tablet containing 5 mg or 20 mg of substrate. Directions for use:Deposit one tablet of the pNPP substrate in a laboratory flask or beaker placed on a magnetic stirrer. Add deionized water in the appropriate amount to reach the required concentration for the assay. Stir until full dissolution and the substrate solution is ready to use.

Substrate for horseradish peroxidase detection assays.

4-Chloro-1-naphtol is peroxidase substrate suitable for use in immunoblotting. The endproduct of the enzymatic reaction is an insoluble blue precipitate that can be observed visually. The precipitate can is not removed by washing with water.

pNPh-b-D-GlcNAc - CAS: [3459-18-5] - C14H18N2O8

pNPh-a-L-Fuc, CAS: [22153-71-5] - C12H15NO7

pNPh-a-D-Man, CAS: [10357-27-4] - C12H15NO8

pNPh-b-D-Xyl - CAS: [2001-96-9] - C11H13NO7

Acetyl Coenzyme A (Ac CoA) is the end product of glycolysis and takes part in the Ac CoA pathway, which is a metabolic pathway for carbon compounds. Ac-CoA is important in cholesterol synthesis. Fatty acids are relatively unreactive and must therefore be activated before reactions with coenzyme A. The key enzyme is the Acyl CoA synthetase, also called thiokinase. The fatty acid is first formed by the thiokinase (attachment of AMP) to acyl adenylate. Acyl adenylate reacts under the cleavage of AMP and the addition of CoA (catalyzed by the thiokinase) to form Acyl CoA. Accordingly, acetic acid can also be activated and forms Acetyl CoA. Concentration and stabilityWorking solution: Optimal solvent: water or aqueous solution with a weak acidic pH (4 to 5).Storage conditions of the working solution: -15°C to -25°C.A solution of 50mg/mL in phosphate buffer, pH7, is stable for 3 weeks at -15°C to -25°C. Unfrozen solutions should be used immediately.

BCIP (5-bromo-4-chloro-3-indolyl phosphate, p-toluidine salt) is a chromogenic substrate for alkaline phosphatase, used in combination with the oxidant NBT (nitro blue tetrazolium) to enhance blue color development. Applications: When alkaline phosphatase conjµgates are used for detection, BCIP is recommended for the following applications: Southern blotting, Western blotting, Northern blotting and dot/slot blots. It is recommended to prepare a 50mg/mL stock solution in DMF.Protocol for Preparation of BCIP/NBT Substrate Solution:Add the following components- 10mL of the Alkaline Phosphatase (AP) buffer (100mM Tris-HCl (pH9.5), 100mM NaCl and 10mM MgCl2)- 33µL BCIP (50mg/mL in DMF) - 44µL NBT (75mg/mL in 70% DMF). NOTE: Avoid exposure to light - Prepare fresh developing solution and use within an hour - BCIP and NBT solutions in DMF do not freeze. They are stable for approximately 2 years when stored at -20°C in the dark.

5-Bromo-3-indolyl-β-D-galactopyranosid (Bluo-Gal) ist used as a substrat of β-Galactosidase. It is a chromogenic substrate suitable for identification of lacZ+ bacterial colonies. Bluo-Gal is designed to replace X-Gal in blue-white selection of recombinant bacterial colonies with the lac+ phenotype. PrincipleWhen Bluo-Gal is hydrolyzed by β-galactosidase, the resulting product will form a blue precipitate. Lac+ colonies grown in the presence of Bluo-Gal turn a deeper blue color more intense than X-Gal, allowing for easy differentiation between lac+ and lac- colonies.

Coenzyme A (CoA, CoASH, HSCoA) is a coenzyme that facilitates enzymatic acyl-group transfer reactions and supports the synthesis and oxidation of fatty acids. CoA is involved in the mechanisms of a wide variety of enzymes.

Glucose 6-phosphate (also known as Robison ester) is glucose sugar phosphorylated on carbon 6. This compound is very common in cells as the vast majority of glucose entering a cell will become phosphorylated in this way. Within a cell, glucose 6-phosphate is produced by phosphorylation of glucose on the sixth carbon. This is catalyzed by the enzyme hexokinase in most cells, and, in higher animals, glucokinase in certain cells, most notably liver cells. One molecule of ATP is consumed in this reaction.

IPTG is commonly used in cloning procedures that require induction of β-galactosidase activity. It is used together with X-Gal (5-bromo-4-chloro-3-indolyl-ß-D-galactoside) or Bluo-Gal (5-Bromo-3-indolyl-beta-D-galactopyranoside) in blue-white selection of recombinant bacterial colonies that induce expression of the lac operon in Escherichia coli. Mode of operation:While IPTG binds to the lacI repressor it is altering the conformation of the lacl repressor, which prevents the repression of the β-galactosidase coding gene lacZ. Thus IPTG is an inducer of the lac operon in bacteria for detection of lac+ colonies. Dissolve in water to e.g. 200mg/mL, sterilise by filtration and store in aliquots at -20°C. For detection of transformants, use in final concentration of 0.1mM up to 1mM in LB-Medium.

IPTG is commonly used in cloning procedures that require induction of β-galactosidase activity. It is used together with X-Gal (5-bromo-4-chloro-3-indolyl-ß-D-galactoside) or Bluo-Gal (5-Bromo-3-indolyl-beta-D-galactopyranoside) in blue-white selection of recombinant bacterial colonies that induce expression of the lac operon in Escherichia coli. Mode of operation:While IPTG binds to the lacI repressor it is altering the conformation of the lacl repressor, which prevents the repression of the β-galactosidase coding gene lacZ. Thus IPTG is an inducer of the lac operon in bacteria for detection of lac+ colonies. Dissolve in water to e.g. 200mg/mL, sterilise by filtration and store in aliquots at -20°C. For detection of transformants, use in final concentration of 0.1mM up to 1mM in LB-Medium.

Ready-to-use 100mM IPTG solution (Isopropyl-ß-D-thiogalactopyranoside) for molecular biology research. IPTG is commonly used in cloning procedures that require induction of β-galactosidase activity. It is used together with X-Gal (5-bromo-4-chloro-3-indolyl-ß-D-galactoside) or Bluo-Gal (5-Bromo-3-indolyl-beta-D-galactopyranoside) in blue-white selection of recombinant bacterial colonies that induce expression of the lac operon in Escherichia coli. While IPTG binds to the lacI repressor it is altering the conformation of the lacl repressor, which prevents the repression of the β-galactosidase coding gene lacZ. Thus IPTG is an inducer of the lac operon in bacteria for detection of lac+ colonies. Shipped as 10 x 1mL solution.

D-Luciferin also named Firefly Luciferin is the most popular and versatile bioluminescent substrate. Firefly luciferase produces light by the ATP-dependent oxidation (Mg2+ as cofactor) of luciferin. It emits a characteristic yellow-green emission in the presence of oxygen, which shifts to red light in vivo. The 560 nm chemiluminescence from this reaction peaks within seconds, with light output that is proportional to luciferase activity when luciferin and ATP are present in excess. Firefly luciferase has long been conjugated to antibodies and used as a label in immunoassays using luciferin as the substrate for detection. One particular advantage to the enzyme is that there is low endogenous luciferase activity in mammalian tissues besides its high sensitivity. Through the utilization of ATP, the reaction can be further used to indicate the presence of energy or life in order to function as a life-death stain. Luciferin is a common reagent used throughout the biotechnology field and specifically for in vivo imaging. Luciferase labeled tumor cells, stem cells or infectious diseases are often inoculated into research animals such as rats or mice for investigation. The injection of luciferin allows for the real-time, non-invasive monitoring of disease progression and/or drug efficacy in these model systems through Bioluminescence Imaging (BLI). Luciferin is also commonly used for in vitro research, including luciferase and ATP assays, gene reporter assays, high throughput sequencing and various contamination assays (e.g., determine cell viability and bacteria counting).

Nicotinamide adenine dinucleotide phosphate reduced - Tetrasodium salt in short NADPH is the reduced version of NADP+ and can transfer a hydride atom to another reaction partner in a redox reaction. The abbreviations NADP+ for the oxidized form, NADPH for the reduced form and NADP in general are proposed by IUPAC. β-Nicotinamide adenine dinucleotide 2'-phosphate (NADP +) and β-nicotinamide adenine dinucleotide 2'-phosphate, reduced (NADPH), comprise a coenzyme redox couple (NADP+ : NADPH) involved in a variety of enzyme-catalyzed oxidation-reduction reactions. The NADP+ / NADPH redox couple facilitates electron transfer in anabolic reactions such as lipid and cholesterol biosynthesis and fatty acyl chain extension. The NADP+ / NADPH redox system is used in a variety of antioxidant mechanisms to prevent the accumulation of reactive oxidation products. NADPH is generated in vivo by the pentose phosphate pathway (PPP).

3-Hydroxy-2-naphthoic-o-toluidide chloroacetate. This product is a histochemical substrate for esterases with improved stability.

Naphthol-AS-BI-phosphate (6-Bromo-2-phosphohydroxy-3-naphthoic acid o-anisidide) is used as histochemical substrate for alkaline phosphatase or as substrate for the fluorometric assay of acid and alkaline phosphatases.

(Nitrotetrazolium Blue chloride, Ditetrazolium dye, Nitro BT). A ditetrazolium compound for demonstrating succinic dehydrogenase activity or glucose-6-phophate dehydrogenase or xanthine oxidase. Used in conjunction with BCIP for detection of alkaline phosphatase. (Negi D.S. and Stephens R.J. (1977) J. Histochem. Cytochem., 25, 149, Vila V. et al. (1984) Clin. Chim. Acta, 138, 215.

Spermine, a polyamine, is derived from spermidine and is required for cell growth in eukaryotes. It acts as regulator of gene expression, inhibits DNA damage and protects the cells from damage by acting as free radical scavenger. In proteomics studies, spermine mediates rapid crystallization of DNA-binding proteins. Spermine is a natural occuring polyamin, which is found in human sperm and other body fluids with a characteristic odour. Spermine is a polyamine involved in cellular metabolism found in all eukaryotic cells. The precursor for synthesis of spermine is the amino acid ornithine. It is found in a wide variety of organisms and tissues and is an essential growth factor in some bacteria. It is found as a polycation at physiological pH. Spermine is associated with nucleic acids and is thought to stabilize helical structure, particularly in viruses. Additionally it is used as starting material for the synthesis of medical products. (Source: Wikipedia: https://en.wikipedia.org/wiki/Spermine).

Spermidine inhibits neuronal nitric oxide synthase (nNOS). As it bind to DNA it might be used as precipitation agent for DNA-binding proteins. Spermidine stimulates T4 polynucleotide kinase activity. As Spermidine and Spermidine hydrochloride deaminates over time, solutions should be sterile-filtered, not autoclaved, if sterile solution is necessary. For the same reason prepare new solutions frequently.

Mitochondriale Dehydrogenasen lebender Zellen sind in der Lage das MTT-Tetrazoliumsalz durch Spaltung des Tetrazoliumringes in MTT-Formazan umzuwandeln. Die Spaltung des Farbstoffes ist direkt proportional zur Zellzahl, steigt aber nicht-linear mit der Funktion der Zeit. Durch MTT kann die Zellaktivierung unabhängig von der Proliferation gemessen werden. Verschiedene Autoren hatten technische Probleme, die zu großen Abweichungen geführt haben. Durch systematische Untersuchung der verschiedenen Parameter konnten diese Fehlerquellen beseitigt werden. MTT = 3-(4,5-Dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazoliumbromid.

3, 3’, 5, 5’-Tetramethylbenzidine (TMB) substrate is a ready-to-use liquid substrate for HRP, specially developed for ELISA procedures. EC-Blue Enhanced™ Substrate turns deep blue (620-650 nm) in the presence of a peroxidase-labelled conjugate. Colour development can be stopped by adding an equal volume of 0.5M H2SO4, which turns the substrate yellow (450 nm). One bottle of EC Blue consists of a solution containing proprietary ready-mixed chromogen substrate reagents, pH3.8. The solution has a straw-yellow colour at room temperature and a light greenish tint when stored at 4°C. The colour returns to normal when warmed overnight to room temperature. Light sensitive! Sensitive against some metal ions! Don't freeze!

X-Gal (5-Brom-4-chlor-3-indolyl-β-D-galactopyranosid) is a chromogenic substrate for β-galactosidase. It produces a blue color that can easily be detected visually. X-Gal is used in the identification of lacZ + bacteria, especially for the detection of ß-galactosidase, which is expressed by the recombinant vectors. X-Gal is often used in conjunction with IPTG (for the induction of the lac promoter (lac operon)), for the screening of blue colonies. It is also used for the screening of ß-galactosidase reporter gene activity in transfections of eukaryotic cells. X-Gal is a substrate of ß-galactosidase, which releases by cleavage of the substrate 5-bromo-4-chloro-3-indoxyl (X). The enzymatically cleaved indoxyl is oxidized to the insoluble indigo. As a stock solution, for example, a 20mg/mL X-Gal solution in dimethylformamide can be prepared. For the use in LB / Amp - plates this stock solution is to be diluted 1 : 1000 (final concentration 20μg/mL) or 1: 500 (final concentration 40μg/mL). The heated medium should thereby be cooled to about 50°C. The amount of X-Gal may be reduced if an X-Gal solution in DMF is sprayed to the agar plates (already with bacterial colonies!). X-Gal is the substrate of choice for blue-white selection of recombinant bacterial colonies with the lac+ genotype.

Chromogenic substrate for beta-glucoronidase. The substance is in use since 1960 in plant tissue sections. Because of the localised colour which X-gluc produces upon reduction, it is extremely useful inidentifying gene presence within most cell types. Recent applications of this product include its use in rapid detection of GUS (E.coli beta-glucoronidase) gene fusion marker in plants. X-gluc has also been reported applications in detection of E.coli in foood samples such as meat, dairy products and shelfish.

X-alpha-D-Gal is a chromogenic substrate for alpha-galactosidase yielding a blue precipitate (X-alpha-Gal is a chromogenic substrate for yeast galactosidase (MEL1) and is used for detecting GAL4-based yeast two-hybrid interactions directly on agar). Used for species differentiation within the family Enterobacteriaceae and differentiation of Bifidobacteria species from Lactobacillus species (see references). Synonmys: 5-Bromo-4-chloro-3-indolyl a-D-galactopyranoside, X-α-Gal; X-alpha-Gal. Other products that are used as a substrate for alpha or beta-Galactosidase: M3225: 5-Bromo-3-indolyl-beta-D-galactopyranoside >M3204: 5-Bromo-4-chloro-3-indolyl-β-D-galactopyranoside (X-Gal) >M3205: 5-Bromo-4-chloro-3-indolyl-ß-D-glucopyranoside (X-Glu / X-Glc) > References: 1. Aho S., Arffman A., Pummi T., Uitto J., A novel reporter gene MEL1 for the yeast two-hybrid system. Anal Biochem. 1997, 253, 270-2. 2. Chevalier P., et al., X-alpha-Gal-base medium for simultaneous enumeration of bifidobacteria and lactic acid bacteria in milk. J. Microbiol. Methods 1991, 13, 75. 3. Gossrau R., Lojda Z., Histochemical detection of alpha-D-galactosidase with 5-Br-4-Cl-3-indoxyl alpha-D-galactoside. Acta Histochem. 1989, 85, 213-22. 4. Perry J.D., Ford M., Taylor J., Jones A.L., Freeman R., Gould F.K., ABC medium, a new chromogenic agar for selective isolation of Salmonella spp. J. Clin. Microbiol. 1999, 37, 766-8.

Rose-Gal, also known as Red-Gal or Salmon-Gal, (6-Chloro-3-indolyl-ß-D-galactopyranoside), is a is a beta-galactosidase substrate which produces a salmon pink colour. Rose-Gal (Red-Gal / Salmon-Gal) serves as an alternative to X-Gal > for the enzymatic detection of Lac+ beta-Galactosidase activity in bacterial colonies. In colorimetric assays, recombinants (white) can be distinguished from non-recombinants (salmon). In conjunction with X-Gluc >, Rose-Gal (Red-Gal, Salmon-Gal) can be used for the simultaneous detection of GUS and Lac + activity on the same plate.

Nicotinamide adenine dinculeotide, min. 95% (HPLC). Suitable for cell culture.Synonym: β-DPN; β-NAD, oxidised form; Coenzyme 1; Cozymase; DPN; Diphosphopyridine nucleotide; Nadide; b-Nicotinamide adenine dinucleotide; b-NAD. Used, among other things, for oxidation of malate. Composition of the measuring buffer for the oxidation of malate: 100mM Tris HCl (pH8), 10mM L-Malat, 0,5mM NAD, 20mM KCl and 1mM MnCl2.

Synonym: β-DPNH; DPNH; Diphosphopyridine nucleotide, reduced form; beta-NADH disodium salt hydrate, reduced. Substrate for the reduction of oxaloacetate and for the reduction of pyruvate Composition of the buffer for the reduction of oxaloacetate: 30mM potassium phosphate pH7.5, 0.5mM oxaloacetate, 0.1mM NADH, 20mM KCl, 5mM MgCl2.Composition of the buffer for the reduction of pyruvate: 10 mM MOPS pH6.1, 10mM Na-pyruvate, 100mM Na2CO3, 0.1mM NADH, 5mM MgCl2. Also available from Genaxxon: M6058 NADPH tetrasodium salt, min. 98% >.

Nicotinamide adenine dinucleotide phosphate - sodium salt in short also NADP as general name for NADP+ and NADPH (IUPAC suggests the abbreviations NADP+ for the oxidized form, NADPH for the reduced form and NADP as a general name). β-Nicotinamide adenine dinucleotide 2'-phosphate (NADP+) and β-nicotinamide adenine dinucleotide 2'-phosphate, reduced (NADPH), comprise a coenzyme redox couple (NADP+ : NADPH) involved in a variety of enzyme-catalyzed oxidation-reduction reactions. The NADP+ / NADPH redox couple facilitates electron transfer in anabolic reactions such as lipid and cholesterol biosynthesis and fatty acyl chain extension. The NADP+ / NADPH redox system is used in a variety of antioxidant mechanisms to prevent the accumulation of reactive oxidation products. NADPH is generated in vivo by the pentose phosphate pathway (PPP). Aqueous solution stable for about 4 weeks at +2°C to +8°C (pH: 2 - 6).

Collagenase-Chromophore-Substrate Component A also named 4-Phenylazobenzyloxycarbonyl-Pro-Leu-Gly-Pro-D-Arg-OH x 2H2O.