Hansatech O2 Electrode |
This won't be a complete description right now.
follow the instructions
follow the instructions
we haven't been entering the atmospheric pressure
Run the water bath with the cooler on until it is stable at the desired temperature. Then turn the cooler off. Watch and record the temperature in the bath. You need to do the run with as little temperature variation as possible, ideally less than 0.1 C.
We are using a Kodak Carosel projector with slides that were computer generated to give grey scales of between 10% and 97%. When these are developed, you need to tell the people not to throw them away. They look light errors in processing and will be automatically discarded.
These slides were calibrated using a Licor Quantum meter. This was done at the same distance from the lense as the chamber. The Hansatech Light meter would be superior, but it is expensive.
Full light is provided by a slide cover with no film.
Dark is with the projector with no slide (the shutter doesn't open) or the lamp turned off.
Our normal routine is to put the culture into the cuvette and let it go for 6 minutes in the dark. Then we employ several different light regimes.
The light regime should be designed to be:
- short (not stress the cells)
- include multiple exposures to each light level
- have long enough exposure at each light that a good reading can be made (about 1 minute)
- keep the cells in a near ambient range of O2 (dark intervals help with this)
- have sufficient values to estimate alpha, Pmax and beta
You can see that these criteria are at odds with each other. In practice, we have been trying to get good estimates of Pmax, alpha and dark_respiation
An example regime is:
START TIME SLIDE % grey LIGHT
uEm-2s-1
0 dark 6 50 285 7 90 26 8 80 71 9 50 285 10 d 0 11 f 1800 12 95 9.4 13 10 842 14 20 530 15 50 285 16 d 0 17 end