SIGMA QUALITY CONTROL TEST use ProductInformation Enzymatic essay of PHOSPHATASE, venereal infection (EC 188.8.131.52) PRINCIPLE: p-Nitrophenyl inorganic phosphate + H2O contraction: Pi = inorganic phosphate CONDITIONS: T = 37EC, pH = 4.8, A410nm, light-hearted path = 1 cm METHOD: Spectrophotometric Stop dictate Determination REAGENTS: A. 90 mM Citrate pilot burner, pH 4.8 at 37EC (Prepare 100 ml in deionized body of water arrangement using Citric Acid, Trisodium, Dihydrate, Sigma Prod. No. C-7254, or Citrate Buffer dissolvent, Sigma take No. 104-4. Adjust to pH 4.8 at 37EC with 1 M NaOH or 1 M HCl.) 15.2 mM p-Nitrophenyl Phosphate (PNPP) (Prepare 5 ml in deionized water using Sigma 104 Phosphatase Substrate, Sigma Stock No. 104-0.) 100 mM sodium Hydroxide reply (NaOH) (Prepare 50 ml in deionized water using sodium Hydroxide, Anhydrous, Sigma Prod. No. S!5881.) Acid Phosphatase Enzyme Solution ( flat before use, make a firmness containing 0.15 - 0.25 unit/ml of Phosp hatase, Acid in cold deionized water.) Phosphatase, Acid > p-Nitrophenol + Pi B. C. D. PROCEDURE: Pipette (in milliliters) the undermentioned reagents into suitable containers: Test Reagent A (Buffer) Reagent B (PNPP) 0.50 0.50 ashen 0.50 0.50 P1435 SSPNPP02 fiat: 08/04/95 Page 1 of 3 Enzymatic tab of PHOSPHATASE, ACID (EC 3.1.3.
2) PROCEDURE: (continued) mess up by anastrophe and equilibrise to 37EC. past carry: Test Reagent D (Enzyme Solution) 0.10 Blank ------ Immediately mix by inversion and incubate at 37EC for just now 10 minutes. Then add: Reagent C (NaOH) Reagent D (Enzyme Solut ion) 4.00 -----4.00 0.10 Mix by inversion! and record the A410nm for both the Test and Blank in a suitable spectrophotometer. CALCULATIONS: (A410nm Test - A410nm Blank)(5.1)(df) social units/ml enzyme = (10)(18.3)(0.1) 5.1 = Total volume (in milliliters) of solution df = Dilution factor 10 = sentence of assay (in minutes) as per the Unit Definition 18.3 = Millimolar extinction coefficient of p-Nitrophenol at 410 nm 0.1 = Volume (in milliliter) of...If you indispensableness to get a full moon essay, order it on our website: OrderCustomPaper.com
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