New ice lithography method enables creation of nanoscale patterns on living tardigrades, advancing integration of biological systems with technology for sensing and monitoring.
(Nanowerk Spotlight) The merging of engineered microscopic patterns with living organisms promises to revolutionize medicine, environmental monitoring, and biological research. Imagine microscopic markers that could track individual cells through development, living organisms modified to visually signal environmental changes, or patterned surfaces that seamlessly connect with biological tissue.
These applications require the ability to create precise patterns and structures on living tissue at the microscopic scale. However, achieving this integration has proven exceptionally challenging due to the fundamental mismatch between biological systems and fabrication methods.
Living organisms operate through complex biochemical processes that require specific temperatures, pH levels, and chemical environments. They rely on carefully balanced ionic concentrations and cannot survive extreme conditions. In contrast, the fabrication of microscopic patterns traditionally demands harsh environments – intense radiation, corrosive chemicals, high temperatures, and vacuum conditions – that destroy biological tissue. This incompatibility has prevented the direct patterning of engineered structures onto living organisms, limiting progress in bio-integration.
Previous attempts to bridge this divide focused on developing patterns that could be attached to organisms after fabrication, or creating biocompatible materials that could interface with living tissue. While these approaches yielded important advances in areas like cell tracking and tissue marking, they could not achieve the precise, direct integration needed for many applications. Soft lithography and bioprinting improved compatibility with biological materials but lacked the nanoscale precision required for advanced patterning.
Researchers also explored various methods to protect biological samples during fabrication, including protective coatings and modified lithography techniques. These efforts produced some success with robust materials like plant cells but failed with more delicate animal tissues. The challenge remained: how to create precise microscopic patterns on living organisms without destroying them in the process.
Scientists at Westlake University have now developed an innovative solution by exploiting the remarkable survival abilities of tardigrades – microscopic animals that can withstand extreme conditions by entering a protective dehydrated state called cryptobiosis. Their research, published in Nano Letters (“Patterning on Living Tardigrades”), demonstrates successful fabrication of nanoscale patterns directly on living tardigrades using a technique called ice lithography, achieving features as small as 72 nanometers while preserving the organisms’ viability.
False-colored SEM image of the tardigrade after rehydration and fixation, with a magnified inset of the blue-boxed area. Scale bar: 10 µm (Image: Reprinted with permission by American Chemical Society)
The technique capitalizes on the tardigrades’ natural resilience by working with them in their cryptobiotic state. After cooling the dehydrated organism to cryogenic temperatures, the researchers coat it with vaporized anisole that forms an amorphous ice layer. A focused electron beam then patterns this ice according to predetermined designs, chemically modifying exposed regions to create stable carbon-rich structures. When warmed, the unexposed ice simply sublimates away, leaving the engineered patterns firmly attached to the tardigrade’s surface. Most remarkably, the patterns remain intact even after the organisms rehydrate and resume normal activity.
The team demonstrated the versatility of their approach by creating various test patterns including nanoscale lines with widths as small as 72 nanometers, precisely arranged microdisk arrays with 3 μm diameters, square patterns with defined 3 μm spacing, and even complex university logos. These “tattoos” showed impressive durability, withstanding stretching, chemical exposure, and repeated dehydration cycles.
The researchers achieved this by developing a specialized carbon nanocomposite substrate that combines carbon nanotubes, graphene microsheets, and a small amount of resin. This innovative material provides both the electrical and thermal conductivity needed for fabrication while its porous structure creates an environment suitable for cryptobiosis. Through careful optimization of process parameters, they achieved a 40% survival rate in patterned tardigrades.
This breakthrough opens new possibilities for integrating engineered structures with biology at the microscopic scale. The ability to fabricate precise patterns directly on living organisms could enable sophisticated biosensors that combine biological processes with visual readouts. It might allow the creation of hybrid systems where living organisms carry engineered markers that enhance their natural capabilities or enable new functions. The technique could potentially extend to other organisms capable of surviving extreme conditions, expanding the toolkit for bio-integration.
Beyond immediate applications, this work demonstrates a novel approach to bridging the gap between biological and engineered systems. By working with organisms in protected states and using carefully controlled fabrication conditions, it may be possible to create more sophisticated bio-hybrid structures than previously imagined. Future developments could lead to living sensors for environmental monitoring, microscopic tracking systems incorporating biological components, or new types of interfaces that merge seamlessly with living tissue.
The research also provides insights into the remarkable resilience of tardigrades, showing that these microscopic animals can survive not only natural extreme conditions but also sophisticated nanofabrication processes. This suggests that other organisms with similar survival mechanisms might be suitable candidates for microscale patterning, potentially expanding the range of living systems that can be enhanced with engineered structures.
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