Tracking Axonal Transport of Organelles in Motor Neurons Using a Microfluidic Device

Begin with a polymer-coated microfluidic chamber with proximal and distal wells connected via multiple microgrooves.

In the proximal well, place mouse embryonic spinal cord tissues rich in developing motor neurons.

Supplement this well with a nutrient medium. Incubate to allow neuron growth and adherence to the polymer-coated surface.

Later, add a growth factor-free nutrient medium to the proximal well.

In distal wells, add a higher volume of nutrient medium containing growth factors to create concentration and volume differences.

These differences cause the axons, the longer neuronal projections, to grow toward the distal wells via the microgrooves.

Add a fluorescent dye solution into both wells.

The dye molecules enter the neurons and stain the mitochondria and acidic organelles at both ends.

In live confocal imaging, fluorescently stained organelles move in both directions between the neural cell bodies and the axons, confirming axonal transport in motor neurons.

Dispose of all medium from the proximal compartment of the MFC. Pick up a single spinal cord explant with a pipette in a total volume of four microliters and inject it as close as possible to the cave. Draw out any exce