Bioimprinting with Milk Protein

Research undertaken at Canterbury University over the past four years by a PhD student is using a milk protein, casein, to create a “template” to 3D-imprint muscle and bone cells. It is hoped this research will have applications in stem cell engineering, re-generative medicine and implantable biomedical devices.

Dr Volker Nock is currently working as a Senior Lecturer in the Electrical and Computer Engineering Department at the University of Canterbury. He is also an Associate Investigator with the McDiarmid Institute for Advanced Materials and Nanotechnology and co-director of the Biomolecular Interactions Centre.

One of Dr Nock’s fields of research is bioimprinting. He has been supervising doctoral candidate, (now Dr) Azadeh Hashemi. Azadeh’s work has built on previous years of work by Professor Maan Alkaisi (also a co-supervisor of Azadeh’s PhD) who developed a method of imprinting the shapes of cells onto plastic.

Casein is one of the major types of protein and is unique to milk. It is often used in food additives and supplements, and is the main component of cheese. 

Azadeh’s research has been focused on making casein-based films with surface patterns, and growing cells on them. She has been able to replicate 3D imprints of cells onto films made of milk protein, which can then be used as a substrate for growing cells. The development of the replication process and control of the biodegradability of the films have been the main focus of her work.

Dr Nock explains bioimprinting is a very unique technique of imprinting a cellular shape into a surface in three-dimension, of every detail you could possibly have. It is a type of casting process, in Azadeh’s case, with a solution of casein being dried on a “master” form.

Azadeh first cultures cells (in the PC2 Lab), in order to get a 3D imprint off them.

A PDMS master mold is then created. A control mold is also made, with regular nano-scale lines, squares and circles, which are used (together with flat surfaces) to compare the results of the cell-shaped imprints. The controls are used to conduct experiments where cell behavior is known, and compare it to the cell behavior on a bio-imprinted surface where the cell behavior is unknown.

The Nanofabrication Lab is where the casein films with bioimprints (or geometric regular features for comparative purposes) are made.

AFM (Atomic Force Microscope) imaging is used to check whether the surfaces created with this process have the shapes and detail of the cells as expected. Because some of the detailed cell structures replicated into the casein are in the nanometer range (a human hair is about 40,000 nanometres in width) there is a need to compliment optical microscopy techniques with atomic force microscopy (AFM) which can resolve smaller detail than a light microscope by scanning an un ultra-sharp (to a few atoms) silicon tip over the surface of interest – such as a casein film.

Azadeh later takes her casein films (that have raised features) back to the PC2 Lab to culture live cells on them.

“We now have a biodegradable, pattern-able surface on which we can culture cells. The patterns can (for example) be used to help guide cells during muscle fibre formation in a Petri dish, while slowly being dissolved by the cells in the process, so that only the finished tissue remains,” Dr Nock says.

The patterns on the films that Azadeh has been able to create mimic the cells’ natural physical environment and can influence cell shape and growth. Potential applications for these micro- and nanostructures are in stem cell engineering, re-generative medicine and implantable biomedical devices.

Dr Nock says Azadeh’s work has shown that shapes of biological cells can be replicated onto casein biopolymers with extremely high resolution. She has also demonstrated that the duration of the degradation of the material can be controlled, and that other cells can be cultured on top of the biopolymer. “One premise is that plastic (bio or not) with the shape of similar cells imprinted on the surface may positively influence the response of other real cells encountering such a surface,” says Volker.

Azadeh says stem cells could be grown on an imprint with patterns of different types of cells to see what type of cell the stem cell would change into. There is a possibility that cancer cells could be stopped from being cancerous by growing them on these patterns and “retraining” or changing them so they behave in a normal, non-malignant way.

There are also potential applications in research, not only in replicating cells to be studied but also providing the material for biodegradable plastic containers in which to carry out operations such as cell cultures.

While the materials have not yet been used in humans, in theory, their application could help recovery from injury or disease with muscle or bone replacement. The films could be implanted where needed to help the cells grow into muscles, bones or other tissues, using the surface patterns as a guide. The biodegradable implant would then dissolve, eliminating the need for secondary surgery to take the implant out.

Azadeh’s work, building on the work of Dr Nock, Professor Maan Alkasai and others, has created exciting possibilities and potentially remarkable uses for this component of milk.