4 replies to this topic

[StevenRed]

I would like to increase the concentration of PSN in my primary astrocyte culture from 1.5% to 2%
But some of my colleagues said it would inhibit cell growth or have some adverse effects...

Just want to ask if this concentration of antibiotic would affect the cultures?

[leelee]

Firstly, why do you want to increase the concentration?

We typically don't use any antibiotics in our cultures at all, you shouldn't really need it.

[StevenRed]

I know it is better not to use antibiotics

In fact, I am using 1.5% of antibiotic dilute in medium to do the subculture of primary astrocyte
When I transfer the cell to 6 well plate for treatment, I use 1% of antibiotic instead

But contamination occur... while my cells in 75cm flask (in 1.5% antibiotic) didn't have such problem
So I decide to use same concentration or even higher in treatment...

[rhombus]

StevenRed, on 13 February 2012 - 07:55 PM, said:

I would like to increase the concentration of PSN in my primary astrocyte culture from 1.5% to 2%
But some of my colleagues said it would inhibit cell growth or have some adverse effects...

Just want to ask if this concentration of antibiotic would affect the cultures?

Dear Stevenred,

As already stated, the use of antibiotics in primary culture should not be necessary. I used to have to prepare endothelial cells isolated from porcine aorta's which were obtained from the local abattoir. An abattoir is not an aseptic environment so I concentrated on reducing the risk of potential contamination.

I am assuming that you prepare your primary astrocytes form rats or mice. Here is what I would try and do to reduce contamination risk:

Do all the dissection in a class II safety cabinet
Thoroughly wash the animal fur with 70% IMS
Autoclave all dissecting instruments and then keep them in the cabinet (when used) in a pot of 70% IMS
Once excised, wash the brain thoroughly with a PBS solution.......many times.
Keep all cells isolated from different brains in seperate flasks/dishes .....and only pool them after many days in culture.
If you are using enzymes to dissociate the brain, make sure that you make them up and FILTER them through a 0.2uM filter

Hope this helps

Kindest regards

Uncle Rhombus

[StevenRed]

OK~I will try to modify it~

Thank you all~