Difference between revisions of "UW-Stout/Caffeine FA19"
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#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. |
− | |||
+ | {| class="wikitable" | ||
+ | |+ Concentrations of Caffeine to Yeast Cells | ||
+ | |- | ||
+ | ! 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 | ||
+ | | - | ||
+ | |- | ||
− | + | ||
− | + | {| class="wikitable" | |
− | + | |+ Caffeine Concentration in Graph: | |
− | + | |- | |
− | - | + | !Series |
− | + | !Concentration in μM | |
− | + | |- | |
− | + | | 1 | |
− | + | | 50 | |
− | - | + | |- |
− | 1 | + | | 2 |
− | + | | 20 | |
− | 50 | + | |- |
− | + | | 3 | |
− | - | + | | 15 |
− | 2 | + | |- |
− | + | | 4 | |
− | + | | 10 | |
− | + | |- | |
− | - | + | | 5 |
− | + | | 8.0 | |
− | + | |- | |
− | + | | 6 | |
− | + | | 5.0 | |
− | + | |- | |
− | 10 | + | | 7 |
− | + | | 2.5 | |
− | + | |- | |
− | + | | 8 | |
− | + | | 1.0 | |
− | - | + | |- |
− | 5.0 | + | | 9 |
− | + | | 0.50 | |
− | + | |- | |
− | 5 | + | | 10 |
− | - | + | | 0.25 |
− | 8.0 | + | |- |
− | + | | 11 | |
− | + | | 0.10 | |
− | + | |- | |
− | - | + | | 12 |
− | + | | 0.0 | |
− | 50 | + | |} |
− | |||
− | 10 | ||
− | |||
− | |||
− | |||
− | |||
− | |||
− | - | ||
− | |||
− | |||
− | |||
− | |||
− | - | ||
− | |||
− | |||
− | 0 | ||
− | |||
− |
Revision as of 08:18, 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 | - |
Series | Concentration in μM |
---|---|
1 | 50 |
2 | 20 |
3 | 15 |
4 | 10 |
5 | 8.0 |
6 | 5.0 |
7 | 2.5 |
8 | 1.0 |
9 | 0.50 |
10 | 0.25 |
11 | 0.10 |
12 | 0.0 |