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Cost-effective method for stamp fabrication used for protein patterning

Name: Akshada Jayant Khadpekar

Department: Chemical engineering

Program: Ph D. (6th year)

Name of supervisor: Prof Abhijit Majumder

Topic of research: Cost-effective method for stamp fabrication used for protein patterning

Description of research work:

The cells in our body come in different shapes and sizes. This shape and size is necessary for the functionality of cells. For example, a neuron's elongated shape is required to collect signals far away from the brain. To study the effect of shape and size of cells on its growth and behaviour in laboratory, a well-established method called micro contact printing (µCP) is used. In this method, protein is printed on the glass or plastic surface using a stamp fabricated. The cells are grown on these protein islands printed in different shapes and sizes to study cell migration, differentiation and other cellular behaviour.

Currently, to get the protein pattern, we need a stamp of that pattern. Photolithography method is used to prepare the stamps of required pattern by using light for etching the mould. Using the soft polydimethysiloxane (PDMS), a rubber-like material which is initially viscous and gets cross linked at high temperature, is used to prepare the stamps from the moulds. However, preparing a mould using photolithography is expensive and needs expertise. It is also unavailable to institutes with low funding, requiring highly sophisticated equipment and cleanroom facilities.

In our study, we fabricated stamps without using lithography or any sophisticated equipment. We prepared the stamps using syringe needles as moulds and polystyrene beads. The PDMS prepolymer was injected into needles of different inner diameter and allowed to crosslink. These solidified PDMS cylinder were then pulled out and arranged in the desired pattern on the glass coverslip to obtain the stamps. These cylinders can also be used to create a curved structured stamp which is otherwise tricky using traditional lithography method. Another stamps were prepared using polystyrene beads, where the beads in aqueous solution were placed on coverslip and slowly heated such that when the water evaporates the beads come close and form a hexagonal packing. The hexagon pattern of beads is covered with a thin layer of PDMS and then used for printing proteins.

To test the reliability of our stamps, we grew mouse muscle stem cells on the patterned protein on a flat surface. We observed that the cells remained on the protein islands only and were viable. The stamps were reproducible and protein deposition was also uniform similar to stamps prepared using traditional lithography method.

There are several advantages of our method of stamp fabrication over the photolithography. The most significant benefit is the cost of stamps prepared using our method is only ₹350 in contrast to ₹1500 a piece using photolithography. The cost of maintenance of highly sophisticated equipment adds to the price which is not required in our methods. Additionally, owing to the stamps' curvature, the same stamp can be used to obtain the pattern of different sizes by either varying the applied pressure or changing the rigidity of the material on which the protein is patterned. More the applied weight and lesser the rigidity of substrate larger is the contact area and hence protein pattern. This is not possible using traditional stamps as they have flat bottom surface. Lastly, it is challenging to obtain the stamps with curvature using photolithography, which is possible using our method. The only disadvantage is we cannot do nano printing with current method. We got the protein pattern of sizes varying between 50 µm to 250 µm.

To conclude, we developed an affordable and easy to use stamp fabrication method, which can help researchers with poor funding expand their field of research. This new cost-effective method of printing can be used in diagnostics and also understand basic biology. The specific protein patterned can be used to capture the certain cell types from the pool of cells or to detect diseased cells from normal cells. These method can also be used for virus identification by patterning antibody specific to surface viral protein