| Description | Products Content:H666048Component5 mLStorageH666048A2×HyperProbe Mixture5×1 mL -20℃. Avoid freeze/thaw cycle.H666048BddH2O5×1 mL-20℃. Avoid freeze/thaw cycle. Products IntroductionHyperProbe Mixture is a premixed system for real-time fluorescence quantitative PCR by probe Products Content:H666048Component5 mLStorageH666048A2×HyperProbe Mixture5×1 mL -20℃. Avoid freeze/thaw cycle.H666048BddH2O5×1 mL-20℃. Avoid freeze/thaw cycle. Products IntroductionHyperProbe Mixture is a premixed system for real-time fluorescence quantitative PCR by probe method (TaqMan, Molecular Beacon, etc.). The concentration is 2×, which contains a new engineered DNA enzyme, PCR Buffer, dNTPs, Mg2+, enhancers and stabilizers, and it is easy to operate. It is mainly used for the detection of genomic DNA target sequences and cDNA target sequences after RNA reverse transcription.This product contains highly sensitive engineered DNA enzyme, which can effectively reduce the non-specific amplification caused by the non-specific binding of primers and templates or primer dimer at room temperature, and at the same time, greatly improve the detection sensitivity and amplification efficiency, and the activation of the enzyme only needs to be incubated at 95 ℃ for 30 s, which greatly shortens the reaction time of PCR. The optimized PCR buffer system and enzyme mixture effectively inhibit the generation of non-specific products, which can significantly improve the PCR amplification efficiency, stronger fluorescence signal and higher sensitivity. caveat1. Before use, mix gently by turning up and down, avoid foaming as much as possible, and use after brief centrifugation.2. Avoid repeated freezing and thawing of the product, which may degrade the performance of the product. This product can be stored at -20℃ for long term storage and protected from light. If frequent use is required within a short period of time, it can be stored at 2-8℃.3. ROX dye is used to correct the fluorescence signal error generated between the wells of the quantitative PCR instrument, ROX is not contained in this product.If you need to match ROX dyes with your instrument, please contact your local sales office or call CombiSense customer service.4006-222-360. UsageThe following examples are conventional PCR reaction systems and conditions, which should be improved and optimized according to the template, primer structure and target fragment size. PCR reaction systemAttention:(1) Usually, a primer concentration of 0.2 M can give better results, and 0.1-1.0 µM can be used as a reference for setting the range. If the amplification efficiency is not high, the concentration of primer can be increased; if non-specific reaction occurs, the concentration of primer can be decreased to optimize the reaction system.(2) The final concentration of the probe used is related to the fluorescence quantitative PCR instrument used, the type of probe, and the type of fluorescent labeling substance, please refer to the manual of the instrument or the specific requirements for the use of each fluorescent probe to adjust the concentration.3) Usually the amount of DNA template is 10-100 ng of genomic DNA or 1-10 ng of cDNA as a reference, as templates of different speciesThe number of copies of the target gene contained in them varies, and a gradient dilution of the template can be performed to determine the optimal amount of template to use. PCR reaction conditions Attention:(1) The enzyme used in this product is activated by pre-denaturation at 95°C for 30 s. Most of the templates can be deconvoluted well under this condition. Under this condition, most of the templates can be well unchained. For templates with high GC content and complex secondary structure, the pre-denaturation time can be extended to 1 minute, so that the starting template can be fully unchained, and if the high temperature treatment time is too long, it will affect the activity of the enzyme; for simple templates, pre-denaturation can also be used for 20 s, and the optimal pre-denaturation time can be determined according to the template situation.(2) It is recommended to use two-step PCR program, the annealing temperature should be 58-64℃ as the reference range, and the annealing temperature can be increased when non-specific reaction occurs. If you can't get good results due to the use of primers with low Tm value, you can try three-step PCR amplification, and the annealing temperature should be set in the range of 56℃-64℃ as a reference. The annealing and extension times for several common instruments are shown in the following table: 20 s for Roche, BioRad, Agilent, Hongshi, Dongshenglong, etc. 30 s for ABI 7000/7300/7500. The annealing/extension times can be set according to the different types of instruments and templates, please follow the instructions of the instruments. The annealing/extension time can be set according to different models of instruments and templates, please follow the instruction manual of the instrument... Read More | Inquire | Protein Purity>95% by SDS-PAGEExtinction Coeff.A276 nm = 0.456 at 1.0 mg/mLMolecular Weight8,759 Da (single chain)General DescriptionNatural human C4a is prepared by cleavage of human C4 protein by human C1s. It is produced during activation of both the classical and lectin pathways of complementProtein Purity>95% by SDS-PAGEExtinction Coeff.A276 nm = 0.456 at 1.0 mg/mLMolecular Weight8,759 Da (single chain)General DescriptionNatural human C4a is prepared by cleavage of human C4 protein by human C1s. It is produced during activation of both the classical and lectin pathways of complement. C4a is a member of the anaphylatoxin family of three proteins (C3a, C4a and C5a) produced by the activation of complement (Hugli, T.E. et al. (1981)). It is an unglycosylated polypeptidecontaining 77 amino acids with a molecular mass of 8,759 daltons. Many of the biological functions of C4a are similar to those of C3a, but the specific activities are far below those of C3a. C4a activity is so low, in fact, that it was initially thought to be inactive. These measured activities include inducing muscle contraction in the guinea pig ileum test (spasmogenic activity), desensitization of muscle to C3a stimulation suggesting that the same receptor for both C3a and C4a is involved (tachyphylactic activity) and inducing vascular permeability in human skin (Gorski J.P. et al. (1979)). C4a does not show tachyphylactic activity against C5a or chemotactic activity. Removal of the C-terminal arginine by serum carboxypeptidase N destroys all these activities (Meuller-Ortiz, S.L., et al. (2009)). C4a appears to act through the C3a receptor (C3aR) which is a G-protein coupled receptor found widely distributed on peripheral tissues, lymphoid cells (neutrohphils, monocyes, and eosinophils) and in the central nervous system (astrocytes, neurons and glial cells) (Law, S.K.A. and Reid, K.B.M. (1995)). Physical Characteristics & StructureMolecular weight: 8,759 calculated molecular mass. Observed mass (MALDI-TOF) is 8,762 + 9 mass units. pI = 9.0 to 9.5 (Gorski, J.P. et al. (1981))Amino acid sequence (77 amino acids): NVNFQKAINE KLGQYASPTA KRCCQDGVTR LPMMRSCEQR AARVQQPDCR EPFLSCCQFA ESLRKKSRDK GQAGLQRC4a is thought to be structurally very similar to C3a and C5a to which it is homologous. Thus its 3D structure is probably similar to the X-ray-derived crystal structureof C3a (Huber, R. et al. (1980)) and the NMR derived structure of C3a: Nettesheim, D.G. et al. (1988); Murray, I. et al. (1999).FunctionSee General Description above. C4a exhibits much weaker biological activities than C3a and C5a. Its activity in inducing erythema and edema in human skin is 25,000-fold weaker than that of C5a and 100-fold weaker than C3a per nanomole. The spasmogenic activity of C4a is 2000-fold weaker than C5a and 100-fold weaker than that of C3a. Due to these differences the role of C4a in these responses in vivo is thought to be negligible.AssaysTwo well established assays for C4a and C3a functional activities include induction of contraction in the guinea pig ileum and the permeation of a dye such as trypan blue from the vasculature into skin. The anaphylatoxins also induce mast cell degranulation, (measured as histamine release), platelet aggregation, IL-1 release from monocytes and the release of prostaglandins and leukotrienes from many cells and tissues. The other assays used for C3a (Dodds, A.W. and Sim, R.B. (1997)) should also respond to C4a, but few reports have described utilizing these assays with C4a. ELISA kits for the assay of C4a levels (or more correctly C4a desArg levels) in blood and other fluids are sold by several companies. These measurements are useful for detecting complement activation in vivo, but the interpretation of their meaning is complicated by the fact that clearance of the anaphylatoxins is rapid. In vivoFreshly drawn normal human serum contains significant levels of all three anaphylatoxins. Although these may represent the resting concentration in vivo it is difficult to draw or store blood without some complement activation so a true in vivo concentration is difficult to determine. The presence of EDTA and Futhan in the collection tubes can minimize this background (Pfeifer, P.H. et al. (1999)). Full activation of all C4 in blood (600µg/mL) would result in ~3,400 nM C4a (~30 µg/mL). Due to the low biological activity of C4a it could require activation of most of the C4 in a small region to achieve the micromolar C4a concentrations necessary to elicit a response.RegulationC4a levels are regulated by three processes: formation, inactivation and clearance. There are two enzymes that cleave C4 and release C4a: C1s and MASP-2. C4a is “inactivated” by removal of its C-terminal arginine amino acid. The product C4a desArg (or C4a without the C-terminal arginine) is produced by the action of the plasma enzyme carboxypeptidase N (Mueller-Ortiz S.L. et al. (2009)). The inactivation is rapid and most C4a is converted to C4a desArg within minutes of its formation. Inactivated C4a lack measurable biological activity. Because of the large number of cells bearing C3a/C4areceptors (endothelial, immune, smooth muscle, neuronal, etc.) the capture, internalization and digestion of C4a and C4a desArg probably results in its removal from circulation.DeficienciesA deficiency of C4 or a deficiency of all of the enzymes that cleave C4 to generate C4a could result in the absence of C4a. There are no known complete deficiencies of all ofthe C4 cleaving enzymes. Examples of C4 deficient humans and mice exist (Wessels, M.R. et al. (1995)), but the degree to which pathologies associated with C4 deficiency are due to the lack of C4 or the absence of C4a is unclear. DiseasesThere are no known diseases connected to C4a or C4a desArg. Precautions/Toxicity/HazardsThe source of C4a is human serum, therefore appropriate precautions must be observed even though the source was shown by certified tests to be negative for HBsAg, HTLV-I/II, STS, and for antibodies to HCV, HIV-1 and HIV-II.Injection can cause anaphylatic shock which is a generalized circulatory collapse similar to that caused by an allergic reaction.Hazard Code: B WGK Germany 3... Read More | EPOCROSTM is a reactive polymer with an oxazoline group on the side chain and is used as a cross-linking agent for water-based resins. Among the water-based polymers developed to address environmental issues and the increasing use of water-based products due to VOC regulations and desolventing, the EPOCROSTM is a reactive polymer with an oxazoline group on the side chain and is used as a cross-linking agent for water-based resins. Among the water-based polymers developed to address environmental issues and the increasing use of water-based products due to VOC regulations and desolventing, the EPOCROSTM WS series is a “water-soluble type” with the following structure.Features and PropertiesHigher reactivity than water-based epoxy, melamine, blocked isocyanateVOC free (EPOCROS™ WS-300 and EPOCROS™ WS-700)High crosslinking density with a small amount addedOne-pack type with long usage timeImproves water resistance, solvent resistance, heat resistance, and the strength of films, etc.Adhesion-imparting possible to PET, OPP, PVC, etc.Fast dryingLow toxicity (Ames Test: Negative, Primary Skin Irritation Test: No irritation)WS Series Product LineupApplicationsNonwoven fabric bindersPigment printingCoatings (metals, films, leather)Paint and coatings, Primers (plastics, building materials, vehicles)AdhesivesMethodASSAY for Product Code DILW:One unit equals a decrease in absorbance of 1.0 per minute at 25°C at pH 7.5 with 2,6-dichlorophenolindophenol as the chromogen.Reagents0.2 M Tris⋅HCl buffer, pH 7.50.006 M NADH. Prepare fresh daily.0.0012 M Dichlorophenolindophenol (DCPIP) Prepare fresh daily.EnzymePrepare a 10 mg/ml solution of enzyme in 0.2 M Tris⋅HCl, pH 7.5.Dilute further immediately before use to give ΔA/min of 0.15-0.20.ProcedureAdjust spectrophotometer to 600 nm and 25°C.Pipette into cuvettes as follows:Mix quickly and measure the decrease in absorbance at 600 nm for 2-3 minutes.Determine the ΔA/min. from the initial linear portion of the curve. (Use portion of curve from t=0 to t=1 minute; the rate is linear for 1/2 to 1 minute.)Calculation... Read More | Purity:>90%, by SDS-PAGE visualized with Coomassie® Blue Staining.Description: High-mobility group box 1 protein (HMGB1), also known as HMG-1 or amphoterin previously, is a member of the HMGB family consisting of three members, HMGB1, HMGB2, and HMGB3. HMGB1 is a DNA-binding nuclear protein,Purity:>90%, by SDS-PAGE visualized with Coomassie® Blue Staining.Description: High-mobility group box 1 protein (HMGB1), also known as HMG-1 or amphoterin previously, is a member of the HMGB family consisting of three members, HMGB1, HMGB2, and HMGB3. HMGB1 is a DNA-binding nuclear protein, released actively following cytokine stimulation as well as passively during cell death. It is the prototypic damage-associated molecular pattern (DAMP) molecule and has been implicated in several inflammatory disorders. HMGB1 signals via the receptor for advanced glycation end-product (RAGE) and members of the toll-like receptor (TLR) family. The most prominent HMGB1 protein and mRNA expression arthritis are present in pannus regions, where synovial tissue invades articular cartilage and bone. HMGB1 promotes the activity of proteolytic enzymes, and osteoclasts need HMGB1 for functional maturation. As a non-histone nuclear protein, HMGB1 has a dual function. Inside the cell, HMGB1 binds DNA, regulating transcription, and determining chromosomal architecture. Outside the cell, HMGB1 can serve as an alarmin to activate the innate system and mediate a wide range of physiological and pathological responses. Extracellular HMGB1 represents an optimal " necrotic marker" selected by the innate immune system to recognize tissue damage and initiate reparative responses. However, extracellular HMGB1 also acts as a potent pro-inflammatory cytokine that contributes to the pathogenesis of diverse inflammatory and infectious disorders. HMGB1 has been successfully therapeutically targeted in multiple preclinical models of infectious and sterile diseases including arthritis. As shown in studies on patients as well as animal models, HMGB1 can play an important role in the pathogenesis of the rheumatic disease, including rheumatoid arthritis, systemic lupus erythematosus, and polymyositis among others. Besides, enhanced postmyocardial infarction remodeling in type 1 diabetes mellitus was partially mediated by HMGB1 activation... Read More |