| Description | Hydrolyzed creatinine, used in the development and bulk formulation of enzymatic creatinine reagents.Enzymatic propertiesSource: MicroorganismEnzyme Committee No. : EC 3.5.2.10Molecular weight: 29 kDa (SDS-PAGE)Isoelectric point: 5.3Km value: 5.0× 10-2 M (Creatinine),8.0× 10-2 M (Creatine)Hydrolyzed creatinine, used in the development and bulk formulation of enzymatic creatinine reagents.Enzymatic propertiesSource: MicroorganismEnzyme Committee No. : EC 3.5.2.10Molecular weight: 29 kDa (SDS-PAGE)Isoelectric point: 5.3Km value: 5.0× 10-2 M (Creatinine),8.0× 10-2 M (Creatine)Inhibitors: Hg2+, Cu2+, Fe3+Optimal pH: 7.0-8.0 Figure 1 Optimum temperature: 65℃ Figure 2pH stability: pH 5.5-10.0 (25℃, 16 h) Figure 3Thermal stability: stable below 65℃ (pH 8.0, 30 min) Figure 4Stability: -25 ~ -15℃ standing storageMaintain over 90% activity for 12 months Figure 5 Assay method for activity1. Principle 2. Definition of enzyme activityUnit enzyme activity is defined as the amount of enzyme required to catalyze a reaction to produce 1µmol creatine per minute under the following conditions.3. Reagent preparationReagent I: 0.3M potassium phosphate buffer, pH 6.5.Reagent II: 0.1M creatinine solution (1.13g creatinine dissolved in 100mL UP water).Reagent III: 4% Na2CO3 solution (4.0g Na2CO3 dissolved in 100mL UP water).Reagent IV: 2% alpha-naphthol solution (2.0g alpha-naphthol dissolved in 100 mL of 99.5% ethanol).Reagent V: 1.2g NaOH and 3.2g Na2CO3 were dissolved in double steaming water at a constant volume of 100 mL.Reagent VI: 0.05% diacetyl solution (0.05mL diacetyl with water to 100 mL).Enzyme diluent: 5 mM Tris-HCl pH 8.04. Operation procedure4.1. Add 0.1 mL reagent I and 0.8mL reagent II into a 5 mL centrifuge tube.4.2. Water bath at 37℃ for 5 minutes.4.3. Add 0.1 mL of the sample to be tested and incubate at 37℃ for 10 minutes.4.4. Add 2.0mL reagent III to terminate the reaction, remove and place on ice.4.5. Add the following reagents in the new 5 mL centrifuge tube in order:The solution for step 4: 80 µLDouble steaming water: 720 µLReagent IV: 400 µLReagent V: 400 µLReagent VI: 400 µL4.6. After standing at 25℃ for 1 h, add 2 mL of double steaming water to dilute.4.7. Use a spectrophotometer to measure the absorbance at 525 nm.* 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 computing Vt: Total volume of reaction liquid (1.0mL);Vs: Enzyme liquid volume (0.1mL);t: Reaction time (10 minutes);df: Dilution ratio;C: Enzyme concentration (mg/mL);1.0: Optical path length (cm);0.0704: Millimolar absorption coefficient (cm²/µmol) of chromophore at 525nm under standard reaction conditions... Read More | Inquire | Inquire | Inquire | Purity>97% SDS-PAGE and HPLC analyses. FunctionLA-PF4 stimulates DNA synthesis, mitosis, glycolysis, intracellular cAMP accumulation, prostaglandin E2 secretion, and synthesis of hyaluronic acid and sulfated glycosaminoglycan. It also stimulates the formation and secretion of plasminogen Purity>97% SDS-PAGE and HPLC analyses. FunctionLA-PF4 stimulates DNA synthesis, mitosis, glycolysis, intracellular cAMP accumulation, prostaglandin E2 secretion, and synthesis of hyaluronic acid and sulfated glycosaminoglycan. It also stimulates the formation and secretion of plasminogen activator by human synovial cells. NAP-2 is a ligand for CXCR1 and CXCR2, and NAP-2, NAP-2(73), NAP-2(74), NAP-2(1-66), and most potent NAP-2(1-63) are chemoattractants and activators for neutrophils. TC-1 and TC-2 are antibacterial proteins, in vitro released from activated platelet alpha-granules. CTAP-III(1-81) is more potent than CTAP-III desensitize chemokine-induced neutrophil activation.Post-translationalProteolytic removal of residues 1-9 produces the active peptide connective tissue-activating peptide III (CTAP-III) (low-affinity platelet factor IV (LA-PF4)). Proteolytic removal of residues 1-13 produces the active peptide beta-thromboglobulin, which is released from platelets along with platelet factor 4 and platelet-derived growth factor. NAP-2(1-66) is produced by proteolytical processing, probably after secretion by leukocytes other than neutrophils. NAP-2(73) and NAP-2(74) seem not be produced by proteolytical processing of secreted precursors but are released in an active form from platelets... Read More |