FULL PAPER 1902365 (1 of 6) © 2019 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim www.small-journal.com Functional 2D Germanene Fluorescent Coating of Microrobots for Micromachines Multiplexing Tijana Maric, Seyyed Mohsen Beladi-Mousavi, Bahareh Khezri, Jiri Sturala, Muhammad Zafir Mohamad Nasir, Richard D. Webster, Zdene ˇk Sofer, and Martin Pumera* T. Maric, Dr. M. Z. M. Nasir, Prof. R. D. Webster Division of Chemistry & Biological Chemistry School of Physical and Mathematical Sciences Nanyang Technological University 637371 Singapore, Singapore Dr. S. M. Beladi-Mousavi, Dr. B. Khezri, Dr. J. Sturala, Prof. Z. Sofer, Prof. M. Pumera Center for Advanced Functional Nanorobots Department of Inorganic Chemistry Faculty of Chemical Technology University of Chemistry and Technology Prague Technická 5, 166 28 Prague 6, Czech Republic E-mail: martin.pumera@ceitec.vutbr.cz, pumera.research@gmail.com Prof. M. Pumera Department of Chemical and Biomolecular Engineering Yonsei University 50 Yonsei-ro, Seodaemun-gu, Seoul 03722, South Korea Prof. M. Pumera Future Energy and Innovation Laboratory Central European Institute of Technology Brno University of Technology Purkyn ˇova 656/123, Brno CZ-616 00, Czech Republic DOI: 10.1002/smll.201902365 1. Introduction Self-propelled artificial micro and nanode- vices capable of autonomous movement are postulated to revolutionize many appli- cations [1–3] ranging from biomedicine [4,5] to environmental remediation. [6,7] How- ever, one major challenge faced is the real time tracking of individual micro/ nanomachines within the swarms of the machines which have pertinent impor- tance. The use of fluorescence has been viewed as a viable option for rapid, in situ visual observation of micromotors. [8] Researchers have focused their atten- tion on addressing this limitation with increased exploration performed in developing novel fluorescent labels and improving fluorescence detection tech- niques. Recently, magnetically guided microrobots have been fabricated with the incorporation of fluorescent agents for in vivo analysis and imaging. [9,10] Nelson and co-workers successfully tracked and monitored the motion of a swarm of artificial bacterial flagella functionalized by a NIR fluorophore. [10] Wang and co-workers fabricated tuneable “on– off” fluoresceinamine functionalized microrobots for rapid and improved detection of sarin and soman simulants caused by enhanced fluid flow in the system. [11] They also reported the detection of toxins using fluorescent-labeled microrobots within a few minutes. [12] In the field of biomedicine, Mg-based fluorescent labeled micromotor pills have been successfully used for targeted delivery of drugs to specific locations within the mouse by an in vivo technique. [13] Conversely, TiO 2 /Au/ Mg micromotors were used for the destruction of Bacillus Anthracis spores and fluorescent imaging was performed to monitor the performance using syto-9 (green) and propidium iodide dyes (red) as labels. [14] However, these studies focused on single labeling of micromachines that perform similar functions. It would be difficult to monitor multiple microma- chines with differing functions if present in the same mixture. As such, we extended and explored the concept of labeling micromachines with fluorophores of different emission wave- lengths, allowing us to tag and locate micromachines in situ simultaneously according to their functions. A novel and stable material, 2D germanene, [15] was used as the florescent Micromachines are at the forefront of materials research as they are self-propelled, smart autonomous systems capable of acting as an intelligent matter. One of the obstacles the field faces is tracking individual micromachines carrying molecular cargo from the rest of the micromachines. Highly stable fluorescent markers based on chemically modified 2D germanene compounds are developed. Two different 2D germanene derivatives, 4-fluorophenylgermanane (2D-Ph-Ge) and methylgermanane (2D-Me-Ge), exhibit different fluorescence under UV light irradiation (excitation at 365 nm), which allows one particular micromotor to be easily distinguished in a mixture of micromotors. This offers a paradigm shift toward a new approach of multiplex detection of self-propelled micromachines. The utility is demonstrated on a drug delivery system, where micromachines carrying a drug are labeled with 2D-Ph-Ge with blue emission while bare micromachines are labeled by 2D-Me-Ge with red emission. This approach of functional fluorescent labeling will pave the way to multiple simultaneous functionalized micromachines identification in complex environments. Microrobots The ORCID identification number(s) for the author(s) of this article can be found under https://doi.org/10.1002/smll.201902365. Small 2019, 1902365