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Allow the standard to equilibrate for at least 15 minutes before making dilutions. Reconstitute lyophilized standard with 1.0 mL of deionized water to yield a stock standard. Reconstitution: After warming lyophilized standard to room temperature, carefully open vial to avoid loss of material. Assay immediately or store samples at ≤-20☌. Centrifuge for 10 minutes at 1000 x g within 30 minutes of collection. Plasma: Collect plasma using citrate, EDTA, or heparin as anticoagulant. Remove serum and assay immediately or store samples at ≤-20☌. Serum: Use a serum separator tube and allow samples to clot for 30 minutes, then centrifuge for 10 minutes at 1000 x g. Specimen Collection and Handling: Specimens should be clear, non-hemolyzed and non-lipemic.Ĭell culture supernatants: Remove any particulate material by centrifugation and assay immediately or store samples at ≤-20☌. Log-log graph paper or automated data reduction Microplate reader capable of measuring absorbance at 450 nmħ. The BD Pharmingen™ TMB Substrate Reagent Set (Cat. Substrate Solution - Tetramethylbenzidine (TMB) and Hydrogen Peroxide. Freshly prepare or use within 3 days of preparation, stored at 2-8☌.Ĥ. SH30088 (heatinactivated) recommended.įreshly prepare or use within 3 days of preparation, with 2-8☌ storage.ģ. Assay Diluent- PBS* with 10% FBS#, pH 7.0. Freshly prepare or use within 7 days of preparation, stored at 2-8☌.Ģ. Coating Buffer - 0.1 M Sodium Carbonate, pH 9.5 7.13 g NaHCO3, 1.59 g Na2CO3 q.s. No 550534) containing Coating Buffer, Assay Diluent, Substrate Reagents A and B, Stop Solution and 20X Wash Buffer Concentrate is recommended.ġ. Sodium azide inactivates the horseradish peroxidase enzyme. Note: Do not use sodium azide in these preparations. Therefore: 1 µg NIBSC 86/504 human IL-2=1.37 µg BD OptEIA™ human IL-2. The conversion factor for NIBSC material is as follows: NIBSC (86/504) equivalent value (IU/ml) = 0.0096 BD OptEIA™ human IL-2 value (pg/ml). Standard 86/504 was evaluated in this set. The NIBSC/WHO First International natural human IL 2 Reference. Standardization: This immunoassay is calibrated against purified Baculovirus-expressed recombinant human IL-2. To design an immunoassay with different sensitivity and dynamic range, the following parameters can be varied: Capture, Detection Antibody titers, Incubation time, Incubation temperature, Assay Diluent formulation, Buffer pH, ionic strength, protein concentration, Type of substrate, Washing technique (i.e., number of wash repetitions and soak times). Sufficient materials are provided to yield approximately 20 plates of 96-wells if the recommended storage, materials, buffer preparation, and assay procedure are followed as specified in this package.īD OptEIA™ Sets allow flexible assay design to fit individual laboratory needs. The OptEIA™ Set for human interleukin-2 (IL-2) contains the components necessary to develop enzyme-linked immunosorbent assays (ELISA) for natural or recombinant human IL-2 in serum, plasma, and cell culture supernatants.