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Stained Spinal Cord Neuron

Neurofilament-specific antibody stain of spinal cord neuron in culture.
(Lynn Gargan, March, 1996)



1. Summary


 

Experimental System: Murine monolayer networks in culture growing on thin film electrode arrays. 
Array Plates: 5 x 5 cm x 1.2 mm soda lime  barrier glass (1,200Å layer of SiO2) 64 ITO thin film microelectrodes terminating in a 0.8 x 0.8 mm  recording area.
Insulation: Polysiloxane (DC 648) 
(hydrophobic)
Deinsulation: Single laser shots, pulsed nitrogen laser 
(337 nm, 2 µJ/µm2)
Tissue Source:  Mouse Embryo CNS tissue (14 days gestation) dissociated and seeded at 500k on 600 mm2 surface areas. 
Networks : Randomized cell body adhesion; 
Spontaneously Active; 
Minimum Size: 1 mm2 (50 neurons) 
Maximum: 600 mm2 (30k neurons)
Surface Preparation: Oxidation of CH3 groups to OH- groups via flaming (1 s exposure to propane or butane flame). Addition of poly-D-lysine and addition of laminin. 


 


Live Neuron on MMEP

Live neuron near row of electrode craters. Indium-tin oxide conductors are 10 µm wide and 1200 Å thick. Craters in 2 µm thick polysiloxane insulation are generated with single laser pulses.



 
 

2. Culture Survival

SUMMARY OF SURVIVAL TIMES ATTAINED (as of 12/96)


 

NOTE: No specific longevity efforts were made. Data represent normal maintenance of cultures that were not selected for experiments at earlier times. All 'old timers' showed spontaneous activity and were pharmacologically responsive. 

 
Tissue No. of Experiments Maximum Survival Time in vitro Size Range of Somata
spinal cord 792  312 d.i.v. 10 - 45 µm
cortex (frontal) 58 81 d.i.v. ----
cortex (auditory) 56 148 d.i.v 5 - 17 µm
olfactory bulb 67 172 d.i.v. ----
cerebellum 6 ---- ----
hippocampus 4 ---- ----


3. Multichannel Recording Work Station

Multichannel Recording Work Station



 

4. Recording Chambers

Recording Chamber
 


Open Chamber Config.
 
 



 



3. Multi-Microelectrode Plates
(MMEPs)


 
 

MMEP 3B
MMEP version 3B
BasePlate: glass 
5 x 5 cm x 1.1 mm
Conductors: indium tin oxide (ITO) 
1200 Å thick, 10 µm wide
Pattern: 0.8 mm x 0.8 mm recording matrix 
4 rows, 16 columns 
40 µm (center to center) between columns 
200 µm between rows
Contacts: 32 left, 32 right 
5 mm long 
0.75 mm wide 
spacing: 0.75 mm
 


 
 

 

MMEP 4A
MMEP 4A matrix (color)
MMEP version 4
BasePlate: glass 
5 x 5 cm x 1.1 mm
Conductors: indium tin oxide (ITO) 
1200 Å thick, 8.0 µm wide 
24 µm x 24 µm terminal pads
Pattern: equally spaced electrodes 
150 µm center to center 
1.2 mm x 1.2 mm recording matrix
Contacts: 32 left, 32 right 
5 mm long 
0.75 mm wide 
spacing: 0.75 mm
 


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  Last updated: August 11, 1999

email gross@nervous.cnns.unt.edu