| Description | Carboxypeptidase B, also known as peptidyl - L - lysine (L - arginine) hydrolase and arginase, is a metalloprotease that can specifically hydrolyze basic amino acids (lysine, arginine, histidine) at the C - terminus of proteins.This product is a recombinant carboxypeptidase B lyophilized powder Carboxypeptidase B, also known as peptidyl - L - lysine (L - arginine) hydrolase and arginase, is a metalloprotease that can specifically hydrolyze basic amino acids (lysine, arginine, histidine) at the C - terminus of proteins.This product is a recombinant carboxypeptidase B lyophilized powder obtained by isolation and purification from recombinant Pichia pastoris, followed by freeze - drying. It is a biochemical - grade genetic engineering enzyme, free from exogenous virus contamination, and does not contain enzyme inhibitors such as DFP, PMSF, and TLCK. It has the same activity and specificity as natural carboxypeptidase B.Product AdvantagesAnimal - free origin: No animal viruses, no pathogenic substances, no exogenous factor contamination, and high safety.High purity and high specific activity.Production scale is over 1000L.Compliance: Production equipment and production environment meet relevant regulatory requirements and conform to GMP guidelines.Complete quality documents: Relevant regulatory support documents can be provided according to customer needs.Product CharacteristicsAppearance: White or off - white powderEnzyme specific activity: ≥150 U/mg proteinMolecular weight: 36kD ± 3.6kDPurity: ≥90% (SDS - PAGE)Protein content: ≥30%Optimal pH: 7.0 - 9.0Solubility: Soluble in buffer solutionRecommended Usage MethodsPreparation of proteinDissolve the protein in a 25mM Tris - HCl solution with pH 7.6.Preparation of enzyme solutionWeigh the required enzyme powder and dissolve it in 25mM Tris - HCl (pH 7.6) to make the enzyme solution concentration 1 - 10mg/ml.ReactionRecommended enzyme amount: Target protein (mass ratio) = 1:50 - 1:1000, and the optimal pH is 7.0 - 9.0.Product ApplicationsProduction of various proteins or polypeptides, such as insulin and its analogs, beauty peptides, antimicrobial peptides, food flavor peptides, etc.Quality detection of antibodies.Determination of amino acids at the C - terminus of proteins.Storage stabilityThe lyophilized powder can be stably stored for at least 24 months at -15°C or below. After being dissolved in 25 mM Tris-HCl (pH 7.6) and stored at -15°C or below, there is no loss of activity even after 10 cycles of repeated freezing and thawing. It is recommended that the dissolved product be stored at 2–8°C for no more than 7 days, and at -15°C or below for no more than 30 days.Precautions and DisclaimerIts activity is competitively inhibited by arginine and lysine; metal ion chelators (such as EDTA) also have an inhibitory effect on the enzyme activity.This product is hygroscopic and should be equilibrated at room temperature before being taken and used.This product is for research use only and shall not be used for the diagnosis or treatment of animals or humans... Read More | Inquire | Biochemical Test:SDS-PAGE (purity > 80%); Western blot with patient sample.Calculated Isoelectric Point:pH 6.10 | GoldStar Probe Mixture is a premixed system specifically designed for real-time fluorescence quantitative PCR using probe methods (TaqMan, Molecular Beacon, etc.), with a concentration of 2 x, containing GoldStar Taq DNA Polymerase, PCR Buffer, dNTPs, and Mg2+. The operation is simple and convenientGoldStar Probe Mixture is a premixed system specifically designed for real-time fluorescence quantitative PCR using probe methods (TaqMan, Molecular Beacon, etc.), with a concentration of 2 x, containing GoldStar Taq DNA Polymerase, PCR Buffer, dNTPs, and Mg2+. The operation is simple and convenient. Mainly used for detecting genomic DNA target sequences and RNA reverse transcription cDNA target sequences, such as gene expression analysis, copy number analysis, SNP genotype analysis, etc., suitable for fluorescence quantification using different types of probe methods. The GoldStar Taq DNA Polymerase contained in this product is a chemically modified, novel and highly efficient hot start enzyme. It has no polymerase activity at room temperature, effectively avoiding non-specific amplification caused by non-specific binding of primers and templates or primer dimerization at room temperature. The enzyme activation requires incubation at 95 ℃ for 10 minutes. The unique combination of PCR buffer system and hot start enzyme significantly improves the amplification efficiency of PCR, with stronger fluorescence signal and higher sensitivity, which can detect single copy templates. By using this product, a wider linear range can be obtained, resulting in more accurate quantification of the target gene. Suitable for all fluorescence quantitative PCR instruments that do not require ROX as a calibration dye.ROX dye is used to correct the fluorescence signal error between wells in quantitative PCR instruments, and is generally used in Real Time PCR amplification instruments from companies such as ABI and Stratagene. The excitation optical systems of different instruments vary, so the concentration of ROX dye must be matched with the corresponding fluorescence quantitative PCR instrument.Instruments that do not require ROX calibration (CW0932): Roche LightCycle 480, Roche LightCyler 96, Bio rad iCyler iQ, iQ5, CFX96, etc.Instrument requiring Low ROX calibration (CW2625): ABI Prism7500/7500 Fast, QuantStudio ® 3 System, QuantStudio ® 5 System, QuantStudio ® 6 Flex System, QuantStudio ® 7 Flex System, ViiA 7 System, Stratagene Mx3000/Mx3005P, Corbett Rotor Gene 3000, etc.Instruments that require High ROX calibration (CW2626): ABI Prism7000/7300/7700/7900, Eppendorf, ABI Step One/Step One Plus, etc.G665832Component5 mLStorageG665832A2×GoldStar Probe Mixture5×1 mL-20℃. Avoid freeze/thaw cycle.G665832BddH2O5×1 mL -20℃. Avoid freeze/thaw cycle. Notes:1. Before use, please gently mix upside down to avoid foaming, and use after briefly centrifugation.2. Avoid repeated freezing and thawing of this product, as repeated freezing and thawing may cause a decrease in product performance. This product can be stored for a long time at -20 ℃, away from light. If frequent use is required in the short term, it can be stored at 2-8 ℃.Usage:The following are examples of conventional PCR reaction systems and reaction conditions. In practical operation, corresponding improvements and optimizations should be made based on different templates, primer structures, and target fragment sizes.1. PCR reaction system Reagent 50 µl Reaction system Final concentration 2×GoldStar Probe Mixture 25 µl 1 × Forward Primer，10 µM 1 µl 0.2 µM¹⁾ Reverse Primer，10 µM 1 µl 0.2 µM¹⁾ Probe，10 µM 1 µl 0.2 µM²⁾ Template DNA 2 µl³⁾ / 50×Low ROX or High ROX（optional）⁴⁾ 1 µl 1 × ddH2O up to 50 µl / Attention:1) Typically, the primer concentration is 0.2 µ M can achieve good results, ranging from 0.1 to 1.0 µ M serves as a reference for setting the range.2) The concentration of the probe used is related to the fluorescent quantitative PCR instrument used, the type of probe, and the type of fluorescent labeling substance. Please refer to the instrument manual or the specific usage requirements of each fluorescent probe for concentration adjustment during actual use.3) The amount of DNA template is usually based on 10-100 ng genomic DNA or 1-10 ng cDNA as a reference. Due to the different copy numbers of target genes contained in templates of different species, gradient dilution can be applied to the template to determine the optimal template usage.4) The excitation optical systems of different instruments vary, and depending on the instrument used for fluorescence quantification, 50 x Low ROX or 50 x High ROX can be added.2. PCR reaction programAttention! The pre denaturation reaction of this product must be completed at 95 ℃ for 10 minutes!Two step PCR Step Temperature Time / Pre denaturation 95℃ 10 min¹⁾ / Denaturation 95℃ 15 s 35-40 cycles Annealing/Extension ²⁾ 60℃ 1 min 35-40 cycles Attention:1) The hot start enzyme used in this product must be activated under pre denaturation conditions of 95 ℃ and 10 minutes.2) It is recommended to use a two-step PCR reaction program. If good experimental results cannot be obtained due to the use of primers with lower Tm values, a three-step PCR amplification can be attempted. The annealing temperature should be set within the range of 56 ℃ -64 ℃ as a reference... Read More | TEV Protease is the 241 amino acid (aa), 27 kDa catalytic domain of the nuclear inclusion a (NIa) protein encoded by the potyvirus, tobacco etch virus (TEV). It may be used in biotechnology to cleave affinity tags from recombinant proteins, either co-translationally orin vitrofollowing purification.TEV Protease is the 241 amino acid (aa), 27 kDa catalytic domain of the nuclear inclusion a (NIa) protein encoded by the potyvirus, tobacco etch virus (TEV). It may be used in biotechnology to cleave affinity tags from recombinant proteins, either co-translationally orin vitrofollowing purification. Its high specificity and activity at a wide range of pH and ionic strength make TEV Protease more versatile than many other proteases used for the same purpose. Unlike factor Xa, enteropeptidase or thrombin, TEV Protease has not been found to cleave at unintended sites, even when present at a high concentration. TEV Protease is a 3C-type protease that cleaves substrates with a consensus sequence of ENLYFQG. Cleavage occurs between Q and G. Since the final aa remains on the cleaved protein where it could potentially affect structure or function, substitution of a variety of aa have been tested. In order of efficiency, S, A, M, Y, D, N, E, K or L may be effectively used in place of G. Several of the remaining aa may also vary, giving a final consensus sequence of ExxYF(M)Q(E)/G(S, A or others) where aa in parenthesis are alternatives and x is any aa. The autocatalytic site of NIa at S2256 has been mutated to an N for improved stability of the protease.Tobacco Etch Virus Protease is a highly site-specific cysteine protease that is found in the tags from fusion proteins. The optimal temperature for cleavage is 30°C. It is recommended that the cleavage for each fusion protein be optimized by varying the amount of recombinant viral TEV protease, reaction time, or incubation temperature. It can be removed by Ni2+ affinity resin... Read More |