User:Jacob Esenther/Notebook/Chem 571/2014/11/12

Biomaterials Design Lab

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Objective Run all tests on the Lysozyme-CaCl2 dialysis Make more lysozyme colloid solution Set up a dialysis of Lysozyme-CaCl2 with new concentrations Description Colloid Synthesis make 0.04621g HAuCl4 / 50mL H2O make 0.0448g Lysozyme / 25mL H2O make a 30:1 HAuCl4 Lysozyme solution 0.5mL HAuCl4, 4.17mL Lysozyme, and 0.33mL 50mL H2O Heat in an oven over 4 hours at 80°C Its go time - 0.6g Lysozyme against 50mM, 5mM, 500μM, 50μM, 5μM CaCl2 Bradford Analysis 50μL sample 200μL Bradford reagent 4:1 DI Water to Bradford Assay 750μL Tris-NaCl Buffer 50mM Tris + 50mM NaCl UV Protein Analysis 10:1 dilutions Fluorescence Analysis 10:1 dilutions ISC of CaCl2 The probe was allowed to sit in solution before taking measurements, this seemed to yield better results. These measurements may need to be taken again to double check Data Bradford Analysis ' ' y=0.0206x+0.0969 Abs. at 590nM Concentration of Lysozyme (ug/mL) Bradford Blank 0.204 5.199029126 5uM CaCl2 0.46 17.62621359 50uM CaCl2 0.467 17.96601942 500uM CaCl2 0.431 16.2184466 5mM CaCl2 0.466 17.91747573 50mM CaCl2 0.43 16.16990291 Table Displaying Calculated Concentrations based off of a Calibration curve of absorbance against a known concentration of Lysozyme. UV Protein Analysis Fluorescence Analysis ISC of CaCl2 ' ISC - CaCl2 Potential=26.344(Concentration)+40.946 ' ' ' Theoretical Concentration (mM) Potential (mV) log (concentration) Actual Concentration (mM) Samples 50.000 89.7 1.850668084 70.90356695 5.000 65.6 0.93584877 8.62678094 0.500 46 0.191846341 1.555415207 0.050 32.1 -0.335788035 0.461542783 0.005 26.1 -0.563543881 0.273184541 Stocks 50.000 93.8 2.006301245 101.4614922 5.000 65.9 0.947236562 8.855978681 0.500 43.8 0.108335864 1.283322663 0.050 28.5 -0.472441543 0.336944566 0.005 15 -0.984892196 0.103539915 Notes Always discuss methods between groups. It can only help you perfect your procedures.