For diffusion modelling of calcium in the sand alginate gel, we want to obtain a video of calcium chloride solution diffusing as a front to crosslink the alginate on a microscope slide.
Materials & Methods
Materials
100 mM calcium chloride
3wt% alginate solution
Earth sand alginate gel (30% 150µm earth sand, 9% CMC, 3% alginate solution, made on 250610)
Spatula
1 mL syringe
microscope slide + cover slip
light microscope
Methods
To prepare the microscope slide, place 200 μl of the bio-ink (using a 1mL syringe) onto a glass slide and press it with a coverslip. Make sure there are no air bubbles.
Mount the slide onto a black surface and below the microscope
Start the video recording on your camera.
Pipette 200 μl of an aqueous solution of 100 mM calcium chloride. Make sure to do this on ONE side of the coverslip border. Don’t cover the entire sample. The Ca2+ ions will diffuse as a flat front along the hydrogel
Let the slide sit and record for 30 minutes.
Save the video. It will be used to measure the diffusion distance pixel by pixel.
Results
Figure 1. Microscope slide setup with 200 μl of gel and 200 μl of calcium chloride solution.
Figure 2. No diffusion across was captured. Gel is on the right and the calcium chloride solution was added on the left.
Observations
Could not see calcium chloride diffusion under microscope (10x objective)
Waited ~10 min before attempting to visualize diffusion
200 μl of gel seemed like too much or too thick
Try 100 μl next to confirm optimal volume
Try plating the gel on half of the slide to leave room for an equal volume of calcium chloride on the other half other slide (amount of calcium chloride to add should be slightly larger than the volume of gel added)
Microscope images taken using iPhone by hand and so it is likely for actual imaging that a better quality microscope with a computer + red laser will be needed
Summary
Need to optimize the amount of gel on slide vs the volume of calcium chloride on slide to improve quality of measuring diffusion
Need a higher quality microscope to successfully record calcium chloride diffusion into bioink
Need to improve earth sand gel formulation to decrease viscosity as thickness seems to be limiting the calcium chloride diffusion process
