HI. Very common problem. I have been doing that a long time ago. We used Sigma BSA fraction V. 2% is enough, more is better. Of course you can't use some BSA fractions or grades without experiencing what you observed. Adding fatty acids has only a metabolic interest. Anyway perfusing with KHB, your heart is totally ischemic and you have to be aware of that. But sometimes it is still ok to explore some factors or products.

Thank you very much. I am currently using fraction V extract from another company. It is a cheaper grade than some available from Sigma, I'll try a better quality one because I've narrowed it down as the sole problem for experiment failure and other researchers using BSA seem to have no problems with it.

Important point : material cleaning and removal of any trace of cleaning agent. The solution is a secret ;). (send me a mail at : [email protected]).Other point, you can perfuse with standard sterile hospital solutions containing proteins (fews) , dextran, HEStarch...etc. but you have to dilute them correctly and to recalculate all ionic contents (I don't know what is your background but any anesthesiologist could help).

Thank you for your help! We have actually repeated the experiment in the same setup with the same solution without BSA (and corrected Ca2+ of course) and it works as intended, so BSA is definitely the problem. I thought about dextran actually, maintains the oncotic pressure very well, with the disadvantage of not allowing fatty acid conjugation, which we were hoping to implement. I'll compare costs and inconveniences against benefits to make a final decision!

I actually am an anesthesiology resident and we have used blood gas analyzers in the past to calibrate our ions (and re-check solution pO2).

Ok, I understand the situation about fatty acids. If it is ok without BSA, well, you have the culprit. You solution pO2 is surely very high around 650-700 mmHg. Hpwever, as you are in anesthesiology, you surely understand that your solution O2 content is very low. Also check your pCO2.Good luck.

Yes, unfortunately we are not currently able to reliably perfuse erythrocytes for multiple reasons (slaughterhouse access, glass reservoirs in the setup, acquiring N2, pump compatibility...) but may manage to do so in the future. We also considered PFC's but they seem to be suboptimal and expensive/time consuming.

pCO2 has been ok as well, even when recirculating. Thank you very much for your tips!

PFC unless you study them are of very little interest. And most of them are even toxic.If you want to perfuse with erythrocytes, glass could be minimal, use silicon tubing and pediatric oxygenators. Here is a reference : TROUVE R. and NAHAS G.

Cardiac dynamics of the Langendorff perfused heart.Proc Soc Exp Biol Med, 180,1985, pp. 303-311. I have a PDF of the article if you mind.

I would appreciate it! Thank you very much. Please send it to [email protected]

Hi guys,

I am a perfusionist here in the US so this interests me quite a bit. I would love to see how you have set up the perfusion circuit if you don't mind.

Thanks,

Sanjay Patel

Give me your email address and I'll send you the paper describing..

Thank you.

[email protected]

Hi André,

In our lab, we dialyse the albumin so that the large fractions of albumin are removed from the solution, that way, it will not clog the coronary vessels of the heart.

That's a solution also but you have to do it very cleanly. No need to add an extra source of problem ... unless you have no alternate.

Hello! To anyone interested, we reached the conclusion that our problems were most likely caused by embolization of poorly dissolved BSA clumps. We took steps to suspend the protein more carefully:

(https://www.labmanager.com/insights/2017/06/7-tips-for-suspending-bsa-in-solution#.W69w3GhKhO8)

Also, we pre-filter the solution by up to 0.21um and use a large diameter in-line filter when recirculating.

This has solved our problems and we now have a stable preparation when perfusing BSA!

Thank you for everyone's input.