Tasks for October 29
- Dialysis of 0.6 g/L Lysozyme vs HCl using MWCO 20000
Preparation of HCl stock solutions
- Need to prepare HCl stock solution of 75 μM, 100 μM, 250 μM, 500 μM and 5 mM with distilled water.
- The initial stock solution of 5 mM HCl was prepared by diluting 0.5 mL of 0.5 N HCl to 50 mL using distilled water
- 0.5v1=5×10-3(50mL); v1=0.5 mL.
- Made remaining stock solutions from 5 mM stock
Preparation of new 0.6g/L lysozyme stock
- Had to prepare new 0.6 g/L lysozyme since it precipitated out of solution.
- Made 50 mL of 0.6 g/L lysozyme solution by adding 0.0380 g of lysozyme (Theoretical value 0.030 g, but some may have been lost in transfer).
Dialysis of 0.6 g/L Lysozyme vs HCl using MWCO 20000
- Note: 100 μM HCl completed evaporated from well, despite being screwed shut.
Bradford Analysis
- Ran bradford analysis of all protein-containing solutions, that is Lysozyme 1, 2, 3, 4 and 5, and all the HCl solutions.
- Remove 100 μL of solution from each chamber (7 in all) and run Bradford analysis
- Bradford reagent should be diluted 1:3 with 50mM Tris/50mM NaCl
- Recall, 100 μL solution + 200 μL diluted Bradford + 700 μL Tris/NaCl buffer
- PS cuvettes, measuring 400 - 800 nm
pH measurements
| Substance
|
pH measurement
|
| 75 μM HCl (1) |
7.142
|
| Lys 1 |
7.116
|
| Lys 2 |
7.423
|
| 250 μM HCl (3) |
5.707
|
| Lys 3 |
6.158
|
| 500μM HCl (4) |
4.944
|
| Lys 4 |
4.921
|
| 5mM HCl (5) |
2.996
|
| Lys 5 |
3.025
|
|
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