Multichannel Systems MEA Manual - Manual (Page 41)

MEA Manual
41
5.4.5 Coating with Poly-D-Lysine (plus Fibronectin)
This coating method is used, for example, for culturing dissociated suprachiasmatic nucleus (SCN) neurons
(on standard 60MEA200/30). It is very stable and therefore especially useful for long-term cultures.
Materials
 Poly-D-lysine 5 mg / 10 mL (= 0.05 % w/v) stock solution (Sigma-Aldrich, Inc., P7280)
 Fibronectin (BD BioCoat™ Fibronectin Cellware) (BD Biosciences)
Fibronectin solution
 Prepare a stock solution of 25 μg/ml fibronectin in distilled water or PBS (phosphate buffered saline) and
store it at 4 °C.
Poly-D-Lysine plus fibronectin solution
 Prepare a 0.01 % (w/v) poly-D-lysine solution, and add fibronectin 1 : 1 (resulting in a final concentration
of 12.5 μg/ml).
Procedure
1. Pipette 10 μl of the poly-D-lysine plus fibronectin solution onto the recording field. Pipette about 50 μl
of sterile distilled water near the rim of the culture chamber.
2. Incubate for 1 h in an incubator set to 35 °C, 65 % relative humidity, 9 % O
2
, 5 % CO
2
; or 37 °C,
100 % humidity, 5 % CO
2
. To avoid a dry out of the liquid, place the MEA in a big Petri dish with lid on.
3. Rinse 2 x with sterile distilled water.
4. Let MEAs air-dry over night on a sterile workbench (laminar flow) with UV light turned on.
5.4.6 Coating with Fibronectin
Fibronectin is a more biological coating alternative, especially used for heart tissues. The adhesion tends
to be very stable, which allows longer cultivation times.
Materials
 Fibronectin (BD BioCoat™ Fibronectin Cellware) (BD Biosciences)
Fibronectin solution
 Prepare a stock solution of 1 mg/ml fibronectin in distilled water or PBS (phosphate buffered saline) and
store it at 4 °C. The stock solution is diluted with water or PBS to a final concentration of 10 μg/ml before
use.