User:Khyra A. Neal/Notebook/Chem 571/2014/10/01: Difference between revisions
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|style="background-color: #F2F2F2" align="center"| | |style="background-color: #F2F2F2" align="center"|[[File:Report.png|frameless|link={{#sub:{{FULLPAGENAME}}|0|-11}}]][[{{#sub:{{FULLPAGENAME}}|0|-11}}|Main project page]]<br />{{#if:{{#lnpreventry:{{FULLPAGENAME}}}}|[[File:Resultset_previous.png|frameless|link={{#lnpreventry:{{FULLPAGENAME}}}}]][[{{#lnpreventry:{{FULLPAGENAME}}}}{{!}}Previous entry]] }}{{#if:{{#lnnextentry:{{FULLPAGENAME}}}}|[[{{#lnnextentry:{{FULLPAGENAME}}}}{{!}}Next entry]][[File:Resultset_next.png|frameless|link={{#lnnextentry:{{FULLPAGENAME}}}}]]}} | ||
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== | |||
* | ==October 1, 2014== | ||
Today's experiment is a continuation of [http://openwetware.org/wiki/User:Khyra_A._Neal/Notebook/Chem_571/2014/09/30 September 30,2014] lab protocol | |||
===Analysis of Dialysis Solutions=== | |||
==Bradford Analysis== | |||
* Remove 20 μL of solution from each chamber (10 in all) and run Bradford analysis | |||
** Bradford reagent should be diluted 1:3 with 50mM Tris/50mM NaCl | |||
** Dilute to 1 mL by adding 780 μL stock 50 mM Tris/NaCl | |||
** Measure UV-Vis from 400 nm - 800 nm using polystyrene cuvettes. | |||
** Run blank of Tris/NaCl buffer | |||
** Run UV-Vis of undialyzed Lysozyme solution with Bradford reagent | |||
* Transfer remaining dialysis solutions into 20 mL extraction vials. | |||
* Measure Ca<sup>2+</sup> of dialyzed solutions that contain Ca<sup>2+</sup> ion using ISE | |||
** Lysozyme dialyzed in both 50 mM CaCl<sub>2</sub> and 500 μM CaCl2 (a total of 4 solutions) | |||
{| {{table}} | |||
| align="center" style="background:#f0f0f0;"|'''Substance''' | |||
| align="center" style="background:#f0f0f0;"|'''Potential (mV)''' | |||
|- | |||
| 50mM CaCl<sub>2</sub> (4) || 80.8 | |||
|- | |||
| Lysozyme (4) || 80.3 | |||
|- | |||
| 500μM CaCl<sub>2</sub> (3)||28.8 | |||
|- | |||
| Lysozyme (3) || 28.1 | |||
|- | |||
|} | |||
==Fluorescence == | |||
* Dilute each dialyzed lysozyme solution by a factor of 100 | |||
* Transfer 100 mL of dialyzed Lysozyme solution to small volume fluorescence cuvette | |||
* Measure fluorescence from 300 nm-550 nm and excitation at 280 nm | |||
* Clean fluorescence cuvette after each use with 1 N HCl, methanol and HPLC water | |||
==Dialysis== | |||
* Prepare another dialysis chamber using 20,000 g MWCO tubing | |||
* On one side add 1 mL of each: | |||
** HPLC water, 50 mM CaCl<sub>2</sub>, 500 μM CaCl<sub>2</sub>, 0.25 mM HCl, 50 mM NaCl | |||
* On opposite side of well add 1 mL of Lysozyme to the wells directly behind the wells containing HPLC water, 0.25 mM HCl, and 50 mM NaCl | |||
* Add 1 mL of Lysozyme Colloid to the wells directly behind the wells containing 50 mM CaCl<sub>2</sub>, 500 μM CaCl<sub>2</sub>. | |||
* Insert screws to prevent leaving/evaporation | |||
* Place on low speed shaker for 1 week | |||
Latest revision as of 00:22, 27 September 2017
Project name | Main project page Previous entry Next entry | ||||||||||
October 1, 2014Today's experiment is a continuation of September 30,2014 lab protocol Analysis of Dialysis SolutionsBradford Analysis
Fluorescence
Dialysis
|