Results

The results of our design project can be broken up into four main sub-results:

Leakage Results

Do our assembled dishes leak?

Cell Adhesion Results

Do the cultured cells adhere to the bottom of the dish?

Cell Alignment Results

Are the cultured cells aligned properly?

XRD Results

Which substrate is most XRD compatible?

Leakage Results 💦

A sample which leaked after incubation.

Initial leakage testing was performed on 6 different sample types: an adhered and non-adhered sample for 3 different membrane thicknesses. After incubation with cell media, both samples of the medium thickness leaked after they were lifted from the plastic trays they were placed in.

This puts the success rate for this batch at ~67%, which is above average.

It is difficult to determine if the failed samples leaked due to their thickness, or if it was due to chance. Thus, although the execution of this component of our design is simple, the rate of success could be higher to reflect a more reliable procedure.

Cell Adhesion Results 🧫

Adherence testing was performed on two of our samples. In one case, the cells were found to be in focus with the grooves underneath a light microscope, indicating cell adherence. In the second case, the grooves were found to be in focus at a different layer than the cells. This indicated that this sample was assembled incorrectly, with the membrane placed upside down. However, as the cells were all found at the same focus level, this still shows that they have adhered to the flat surface.

This demonstrates that PDMS is a viable choice for cell culturing; however, more samples could have been examined for stronger evidence.

Cultured cells stained with DAPI for better visualization. Micropatterned grooves are also visible as parallel lines.

Cell Alignment Results 📏

To measure cell alignment, our client kindly provided us images of cardiomyocytes from his own work. Compared to our own images, it was preferable to use our client's as they were at a higher resolution and stained appropriately which allowed for the visualization of sarcomere fibers.

ImageJ, an imaging processing program, was used to quantitatively test for cell alignment. More specifically, an analytic tool called Directionality by Local gradient orientation was used. The analysis generates a histogram which plots the amount of structures within the image against their orientation direction in degrees, ranging from -90° to 90° (with 0° indicating a completely horizontal direction). A Gaussian fit is then applied to the most significant peak in the histogram.

The cells grown on flat PDMS show a relatively flat distribution with a slight peak at -63° and a stdev of 27°. The cells grown on micropatterned PDMS shows a distribution with a more narrow peak at 8° and a stdev of 17°.

A lower stdev value indicates that the structures within the image have a preferred orientation direction along the peak angle. Thus, the micropatterned PDMS can be quantitatively shown to improve cell alignment.

XRD Results 🔆

Raw XRD pattern images for each of our substrate types were sent over from Argonne National Lab. In general, the diffraction patterns resembled a ring of light.

To analyze each image, ImageJ was used to generate a photon intensity heat map. A higher photon intensity means that the material lets more photons through (i.e. greater x-ray transparency, which is ideal for XRD).

From the heat map, an intensity histogram was generated. A higher mean value indicates a higher x-ray transparency. After normalizing the mean intensity for each substrate, the Curi Bio polymer was found to be the most transparent, 2 orders of magnitude greater than the other substrates.

Curiously, thicker substrates were found to be more transparent, which is the opposite of what we expected. This result may be due to some scattering phenomenon or material property we are unfamiliar with - more research will have to be done.

Page edited by: Andrew Setiadi

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