| Description | ApplicationIt is used for the development and mass preparation of total bile acid (TBA) reagents.Enzymatic properties (note that 2+, +, 3+ are superscripts)Source: MicroorganismEnzymology Committee Number: EC1.1.1.50Molecular weight: 30 kDa (SDS-PAGE)Isoelectric point: 5.2Km value: 3.6×10-5M (ApplicationIt is used for the development and mass preparation of total bile acid (TBA) reagents.Enzymatic properties (note that 2+, +, 3+ are superscripts)Source: MicroorganismEnzymology Committee Number: EC1.1.1.50Molecular weight: 30 kDa (SDS-PAGE)Isoelectric point: 5.2Km value: 3.6×10-5M (androsterone), 4.7×10-5M (NAD+)Inhibitors: Cu²⁺，Ag⁺，Hg²⁺，Zn²⁺，Fe³⁺ Optimum pH: 8.5-9.5 Figure 1Optimum temperature: 55℃ Figure 2pH stability: 6.0-9.0 (25℃,16h) Figure 3Thermal stability: Stable below 37℃ (pH9.0, 30min) Figure 4Stability: -25 ~ -15℃ standing store for 12 monthsMore than 90% activity Figure 5Protective agent: BSA Assay method for activity1. PrincipleThe NADH produced by the reaction can be detected with a spectrophotometer at 340nm.2. Definition of enzyme activityUnit enzyme activity is defined as the amount of enzyme required to catalyze the production of 1µmolNADH per minute under the following conditions.3. Reagent preparationReagent I: 0.1M sodium pyrophosphate (adjust pH to 8.9 with HCl).Reagent II: Dissolve 319mg NAD+ into 25mL double steaming water, adjust the pH to 7.0-7.5 with solid NaHCO3, and adjust the volume to 30mL with double steaming water.Reagent III: Dissolve 30mg into 100mL methanol.Enzyme diluent: 10mMTris-HCl, pH 9.0.4. Operation procedure1. Add 2.6mL reagent I, 0.2mL reagent II and 0.1mL reagent III into a 3mL colorimetric dish and mix well.2. Preheat the reaction mixture at 25°C for 5min.3. Add 0.1mL enzyme liquid to the reaction mixture, mix it well, react at 25°C, and record the absorbance change within 1min at 340nm with a spectrophotometer (∆As).* Replace enzyme liquid with enzyme diluent, other steps are the same, the absorbance of the resulting solution is blank absorbance (∆Ab)∆A=∆As-∆Ab5. Vitality computing3.00: total volume of reaction liquid (mL);0.10: enzyme liquid volume (mL);1.0: optical path length (cm);df: dilution ratio;C: Enzyme concentration (mg/mL);6.22: Nanomolar absorption coefficient of NADH at 340nm (cm2/µmol)... Read More | Inquire | DescriptionApolipoprotein E (ApoE) is present in the brain and is mainly produced by astrocytes. It is a 299 amino acid glycoprotein of 34kDa. It is present in all classes of lipoproteins except LDL (low-density lipoprotein). APOE gene has three alleles, such as APOE ε3, APOE ε4and APOE &#DescriptionApolipoprotein E (ApoE) is present in the brain and is mainly produced by astrocytes. It is a 299 amino acid glycoprotein of 34kDa. It is present in all classes of lipoproteins except LDL (low-density lipoprotein). APOE gene has three alleles, such as APOE ε3, APOE ε4and APOE ε2. It is located on human chromosome 19q13.Preparation instructionsFormLyophillized from a 0.2 µm filtered solution in 20 mM sodium phosphate, pH 7.8.Principle... Read More | Reverse transcriptases are enzymes encoded in retroviruses viral genome. The enzyme is responsible for transcription of the viral RNA to produce a dsDNA that can be inserted into the host genome.Reverse transcriptases are multifunctional enzymes. These enzymes exhibit an RNA and DNA directed Reverse transcriptases are enzymes encoded in retroviruses viral genome. The enzyme is responsible for transcription of the viral RNA to produce a dsDNA that can be inserted into the host genome.Reverse transcriptases are multifunctional enzymes. These enzymes exhibit an RNA and DNA directed polymerase activity. In addition reverse transcriptases catalyze the degradation of RNA in an RNA-DNA hybrid. The exonucleolytic activity proceeds in a 5' ---> 3' direction. The RNA or DNA directed activity requires a template (RNA or DNA) and a primer. The following is a schematic illustration of the reaction:Unit definition: One unit incorporates 1 nanomole of tritiated dTMP into acid insoluble productsusing poly(A)•oligo(dT) 12-18 as the template-primer in 20 minutes at 37° C.ApplicationsHIV reverse transcriptase is used for research on the AIDS primer. However it can be substituted for AMV reverse transcriptase, which is mainly used to transcribe mRNA into double stranded cDNA, that can be inserted into prokaryotic vectors. The enzyme can also be used with either single stranded DNA or RNA templates to make probes for use in hybridization experiments. It can be used for labeling the termini of DNA fragments with protruding 5' termini. The enzyme can also be used to sequence DNAs by the dideoxy chain termination method of Sanger when the Klenow fragment of E. coli DNA polymerase I, or the T7 DNA polymerase yield unsatisfactory results.Reagents0.05 M Tris, pH 8.3, containing 0.008 M MgCl21 mg/ml polyadenylic acid in water (poly A)DNA primer:Oligo d(T)12-181 µ mole dTTP/mL stock solution[methyl-3H]-Thymidine 5'-triphosphate (3H-dTTP)dTTP-3H-dTTP working mix: Add 1-2 µL 3H-dTTP per mL of 100 nmol/mL dTTP in order to obtain 1 to 1.5 x 105 cpm/mL1% bovine serum albumin10% perchloric acid1% perchloric acidBuffer substrate reaction mixture: Prepare fresh, immediately before use:For each 1mL of reaction mixture required mix:0.7 mL Tris/HCl, pH 8.3, 0.008M MgCl20.3 mL 1 mg/mL poly(A) RNA template0.005 mL 0.02 mg/mL oligo d(T)12-18 DNA primer0.02mL 1% BSAEnzymedilute as needed wtih 0.05M Tris/HCl, pH 8.3, 0.008M MgCl2 containing 0.1 mg/mL (1%) BSAProcedurePipette into each tube as follows:Buffer substrate mix：0.1 mLdTTP-3H3-dTTP：0.1 mLEnzyme：5-10 µLIncubate 20 minutes at 37° C. Stop reaction by adding 1 ml 10% cold perchloric acid. Filter through 0.2µ manifold filters used with Millipore vacuum manifold. Wash four times using 2mL 1% cold perchloric acid/wash. Transfer filter to scintillation vials. Add 2mL Cellosolve (or 2-methoxyethanol) to dissolve filter. Filters become opaque upon addition of Cellosolve. Make sure filters are dissolved before proceeding. Add 10mL scintillation cocktail and count.Calculation... Read More | Purity:>90%, by SDS-PAGE visualized with Coomassie® Blue Staining.Implicated in the control of cell proliferation and cellular aging. May also act as a chaperone |