Difference between revisions of "UW-Stout/Caffeine FA19"
| Line 16: | Line 16: | ||
#Fill the wells with different concentrations of caffeine solution. | #Fill the wells with different concentrations of caffeine solution. | ||
#After the wells are completed, place the 96 Well assay plate into the SpectraMax Plus 384 Microplate Reader. Data will be collected every 5 minutes for the next 24 hours. | #After the wells are completed, place the 96 Well assay plate into the SpectraMax Plus 384 Microplate Reader. Data will be collected every 5 minutes for the next 24 hours. | ||
| − | #Once results are computed the data curve should be put on the growth data curve | + | #Once results are computed the data curve should be put on the growth data curve. |
| − | Volume of the Stock solution of 10mM ( | + | |
| − | 0 | + | {| class="wikitable" |
| − | 50 | + | |+ Concentrations of Caffeine to Yeast Cells |
| − | 50 | + | |- |
| − | - | + | ! Concentration of solution added to yeast (μM) |
| − | 0 | + | ! Volume of solution added to yeast (μl) |
| − | 0.10 | + | ! Water added to the solution (μl) |
| − | 50 | + | ! Volume of the Stock solution of 100mM (μl) |
| − | 49 | + | ! Volume of the Stock solution of 10mM (μl) |
| − | - | + | |- |
| − | + | | 0 | |
| − | 0.25 | + | | 50 |
| − | 50 | + | | 50 |
| − | 47.5 | + | | - |
| − | - | + | | 0 |
| − | 2.5 | + | |- |
| − | 0.50 | + | | 0.10 |
| − | 50 | + | | 50 |
| − | 45 | + | | 49 |
| − | - | + | | - |
| − | 5 | + | | 0 |
| − | 1.0 | + | |- |
| − | 50 | + | | 0.25 |
| − | 40 | + | | 50 |
| − | - | + | | 47.5 |
| − | 10 | + | | - |
| − | 2.5 | + | | 2.5 |
| − | 50 | + | |- |
| − | 47.5 | + | | 0.50 |
| − | 2.5 | + | | 50 |
| − | - | + | | 45 |
| − | 5.0 | + | | - |
| − | 50 | + | | 5 |
| − | 45 | + | |- |
| − | 5 | + | | 1.0 |
| − | - | + | | 50 |
| − | 8.0 | + | | 40 |
| − | 50 | + | | - |
| − | 42 | + | | 10 |
| − | 8 | + | |- |
| − | - | + | | 2.5 |
| − | 10 | + | | 50 |
| − | 50 | + | | 47.5 |
| − | 40 | + | | 2.5 |
| − | 10 | + | | - |
| − | - | + | |- |
| − | 15 | + | | 5.0 |
| − | 50 | + | | 50 |
| − | 35 | + | | 45 |
| − | 15 | + | | 5 |
| − | - | + | | - |
| − | 20 | + | |- |
| − | 50 | + | | 8.0 |
| − | 30 | + | | 50 |
| − | 20 | + | | 42 |
| − | - | + | | 8 |
| − | 50 | + | | - |
| − | 50 | + | |- |
| − | 0 | + | | 10 |
| − | 50 | + | | 50 |
| − | - | + | | 40 |
| + | | 10 | ||
| + | | - | ||
| + | |- | ||
| + | | 15 | ||
| + | | 50 | ||
| + | | 35 | ||
| + | | 15 | ||
| + | | - | ||
| + | |- | ||
| + | | 20 | ||
| + | | 50 | ||
| + | | 30 | ||
| + | | 20 | ||
| + | | - | ||
| + | |- | ||
| + | | 50 | ||
| + | | 50 | ||
| + | | 0 | ||
| + | | 50 | ||
| + | | - | ||
| + | |- | ||
Revision as of 08:05, 5 December 2019
Caffeine Dilution and Concentration of Caffeine Protocol
Materials
- 0.22g Caffeine
- Sterile H2O
- Syringes and purifier
- 96 Well assay plate
Procedure
- Make a 100μM stock solution of caffeine and H2O.
- Take 0.1942 grams of caffeine to 10ml of H2O to form a stock solution of 100C.
- Make another stock solution of 10μM of caffeine and H2O.
- Dilute the 100μM stock solution of 10μM stock solution by adding 4μl of 100μM of the stock solution and 36μl sterile H20.
- Filter the caffeine solution using the syringes and purifier then put in microfuge tubes, freeze at -20°C.
- Dilute the 100μM stock solution into the 12 concentrations using the table below.
- Fill the wells with different concentrations of caffeine solution.
- After the wells are completed, place the 96 Well assay plate into the SpectraMax Plus 384 Microplate Reader. Data will be collected every 5 minutes for the next 24 hours.
- Once results are computed the data curve should be put on the growth data curve.
| Concentration of solution added to yeast (μM) | Volume of solution added to yeast (μl) | Water added to the solution (μl) | Volume of the Stock solution of 100mM (μl) | Volume of the Stock solution of 10mM (μl) |
|---|---|---|---|---|
| 0 | 50 | 50 | - | 0 |
| 0.10 | 50 | 49 | - | 0 |
| 0.25 | 50 | 47.5 | - | 2.5 |
| 0.50 | 50 | 45 | - | 5 |
| 1.0 | 50 | 40 | - | 10 |
| 2.5 | 50 | 47.5 | 2.5 | - |
| 5.0 | 50 | 45 | 5 | - |
| 8.0 | 50 | 42 | 8 | - |
| 10 | 50 | 40 | 10 | - |
| 15 | 50 | 35 | 15 | - |
| 20 | 50 | 30 | 20 | - |
| 50 | 50 | 0 | 50 | - |
