Problem with the hippocampal cell cultures

Dear MCS/MEA users

We have some problems with (E17) hippocampal cell cultures on MEAs (100/10 internal ref). We reused the MEAs several times, but after the 5th times our cultures start to die out in DIV6-7. In the cultivation we didn't observe any attachment problem with the neuronal cells and the MEA surface. We observed that the glial cells aren't growing very well and the hippocampal cells are much more sensitive than it should be. And the network is very aggregated. Does anybody have the same problem?
For cleaning we use 1% Tergazyme solution. For coating we use PLL and 10 ug/ml Laminin.

Thank you very much!

Aniko

Dear Aniko,

sorry to hear about the problems you are experiencing with the cell culture.

Do you use Plasma Cleaner to obtain a more hydrophilic surface?
While a hydrophobic surface should also affect adhesion, it might influence cell
survival as well. Especially as you mention aggregation this would be my first guess...

Does the baseline noise level look ok?

Greetings

Thomas

Dear Thomas,

we only used 1% solution of Terg-a-zyme for cleaning.
I checked the baseline noise level. It looks OK for me.
The voltage axis are the following:
-the first measurement data: approximately -62,72uV between +76,36 uV.
-the last measurment data: approximately -62,74 uV between +66,08 uV.
Do you think it is OK? The punctual types of the chips are 60ThinMEA100/10-ITO.

Greetings

Aniko

Dear Aniko,

the roughly +/- 65uV seem very high to me.
while the 10um diameter electrodes do have a slighly higher impedance and thus a higher noise level, they should still be below 30uV peak-peak noise.

Please consider a plasma treatment to clean and hydrophilize the surface and re-check noise with just buffer.

Greetings

Thomas

Dear Thomas,

We tryed out the plasma treatment but we didn't find the punctual parameters of this. Our parameter was:
-duration 2 min,
-power cirka 100W,
-and we used O2.
Does it look fine for you? What parameters do you recommend?

Thank you for your reply in advance
Aniko

Dear Aniko.
the power is a bit on the upper end, but should be O.K. Time and O2 is fine. Use around 0.2mbar pressure.

Cheers Frank MCS