This article describes a method for fabricating microbeads that encapsulate neurotoxic amyloid-beta cells using a biocompatible polymer. The process mimics the chronic accumulation of amyloid-beta seen in Alzheimer's disease, allowing for controlled release and cell proliferation.
Take a suspension of cells that secrete neurotoxic amyloid-beta mixed with a biocompatible polymer.
Attach the syringe with the suspension to a microbead fabrication device, then push it through as a continuous liquid stream.
A vibration frequency in the device breaks the stream into uniform, micron-sized droplets.
These droplets pass through an electrostatic field and acquire uniform charges to repel each other, preventing merging.
The droplets then fall into a gelation bath containing calcium ions. The ions cross-link the polymer, encapsulating the cells in spherical microbeads.
Incubate the beads to complete gelation, then collect them on a filter.
Invert the filter over a tube, add a culture medium to wash the beads into the tube, and transfer the suspension to a flask containing the medium.
The polymer matrix allows nutrient access to support cell proliferation while enabling the slow release of amyloid-beta, mimicking the chronic accumulation seen in an Alzheimer’s diseased brain.
To fabricate the microbeads, in a 20-milliliters syringe, load five milliliters of the cell alginate suspension and attach a syringe to the encapsulator. To start the encapsulator, activate the flow, which will push the cell alginate suspension through the feeder, and a stream of droplets will be extruded through the nozzle. Collect the first one milliliter in the waste beaker to avoid the initial non-uniform stream. Then, continue to run the remaining four milliliters, allowing the droplets to fall into the calcium chloride gelation bath.
Upon contact with the gelation bath, the alginate in the droplets instantly crosslinks with the calcium ions in the gelation bath, forming spherical microbeads. After one minute, remove the gelation beaker from the magnetic platform and allow the microbeads to rest for a further four minutes without agitation to complete their gelation at room temperature. To retrieve the microbeads, first use a pair of sterile tweezers to remove any large alginate debris or artifacts. After cutting the end of a plastic pipette, use it to transfer the microbeads from the gelation bath to a 74-micrometer mesh filter held over a waste beaker.
To ensure success, we always cut the end of a plastic pipette to avoid damaging the microbeads and we do this every time we transfer beads from one vessel to another after encapsulation.
Invert the mesh filter over a centrifuge tube. Pipette the appropriate culture medium over it to wash the beads down the tube and allow them to equilibrate in that medium for five minutes. Then, transfer them to a flask previously filled with the appropriate medium for incubation and further experiments.
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