How to keep pH of organ perfusate stable? - (Jan/18/2006 )
Hi.....I am a PhD student. I am doing a set of experiment to perfuse some freash isolated animal eyes. I have a problem.
The pH of the perfusate was kept changing by the gas I used. Initially, I adjusted the pH of the perfusate to 7.4 after 30min of carbogen (95% O2 / 5% CO2) gasing. However, after 30-60min, the pH must drops to 7.15-7.25. Then I re-adjust the pH again and again, but finally the pH still dropped to 7.2.
I have also tried to gas it with normal air. But the pH increased to 7.58. Then I adjusted it to 7.4 again, but finally it increased to 7.58.
Does anyone know how to maintain the pH of perfusate, please?
The perfusates I tried include: Krebs', Ringer, Medium 199. But they all suffered by the same problem.
Forrest 
The pH keeps dropping because those eye cells are still alive and respiring... taking up O2 and giving off C02... which causes the pH to drop.
To correct the problem, you need to set up a circulating perfusion whereby you use gas permiable flexible tubing that passes thru the eye, and thence to a 'oxygen bubbler' which allows for exchange of the CO2 for fresh O2 and maintains a neutral pH.
To correct the problem, you need to set up a circulating perfusion whereby you use gas permiable flexible tubing that passes thru the eye, and thence to a 'oxygen bubbler' which allows for exchange of the CO2 for fresh O2 and maintains a neutral pH.
I am sorry for not clearifying my system enough. In my system, the perfusate will not be recycle. They will pass the eye and enter a wast collector.
However, my problem is that, even I just keep the perfusate in room temperature, the pH of the perfusate will change according to the type of gas bubbling it. So, it is just like gasing a cup of medium with pH drop caused by bubbling it with carbogen. Its pH will increase, if I bubble it with normal air.
Is there any measure to keep the pH through the whole period of experiment, please?
Could I continously adjust the pH, please? I afraid that will cause osmolarity problem.
Is it possible to keep a cup of medium, without cells of course, after gasing it with carbogen for certain period of time (e.g 30min), then leave it there without bubbling please?