This study presents the high-precision microinjection of fluorescent micro-gel beads into Caenorhabditis elegans trapped in a suction microchannel. In our previous works, we demonstrated survival microinjection by a conventional micromanipulation technique. However, the focal planes differed between the tip of the microinjection tool and the target axon inside the C. elegans body. To resolve this problem, we here propose a suction microchannel that traps C. elegans during the microinjection operation. The focal plane of the target nerve axon matches that of the fluorescent microbead in the microinjection tool, enabling high-precision microinjection into the interior of the C. elegans body under a microscopic view. In an experimental evaluation, the positioning accuracy of the injection into C. elegans was within the target accuracy (15 μm). The head-flrst navigation alignment of C. elegans along the microchannel was controlled by electrotaxis. Injection of the fluorescent micro-gel bead into the C. elegans body was quantitatively confirmed by confocal microscopy.

This paper presents a novel In situ nanomanipulation integrated with scanning electron microscope- computed tomography (SEM-CT) imaging system for 3D nanomanipulation. In our previous works, a nanorobotic manipulation system was established inside an environmental-SEM (E-SEM) for water-contained samples, including biological organism, based on a real-time high resolution SEM observation. However, the SEM image is limited in two dimensional (2D) and surficial information from the signals of secondly electrons. For nanosurgery applications, such as nanoinjection, it is needed to evaluate the sample in 3D space with its internal information after manipulation. The SEM-CT imaging system is developed for In situ nanomanipulation based on SEM observation. The CT is an effective method to obtain the internal 3D information as a non-destructive manner. The imaging resolution of our SEM-CT system is in less than 400 nm. A Caenorhabditis elegans (C. elegans) was used as a target of biological sample. To improve the contrast of SEM-CT imaging of C. elegans, the X-ray was tested by generating using brass and copper materials. Finally, the nanoinjection was demonstrated with SEM-CT imaging system to C. elegans using the nanoinjector which was fabricated by focused ion beam (FIB) process.

This study presents the survival microinjection into Caenorhabditis elegans (C. elegans) with in vivo observation based on micromanipulation. The microinjections were achieved with micro-gel beads which are enable to encapsulate chemicals for injection. In this study, a fluorescent material was used to evaluate the injection positional precision inside the C. elegans. The fluorescent microbead was picked up at the tip of a micropipette injection tool and injected by a piezo actuated microinjector. The distance between the injected micro-gel bead and closest nerve axon was measured as 20.3 μm and 16.5 μm by in vivo observation of a confocal microscopy. The two types of pipette tools were used to evaluate the success and survival rates of microinjection, and the smaller pipette (pipette A, 0.8 μm in diameter) showed higher rates as 50 % and 67 % respectively.

This paper presents a novel selective nano-injection method using fluorescent nanobeads based on nanomanipulation for in-vivo single cell analysis. The nanomanipulation system was constructed under hybrid microscope. The hybrid microscope consists of the optical microscope (OM) and environmental-scanning electron microscope (E-SEM) to realize biological specimen analysis by optical imaging including fluorescent imaging, and nano-scale manipulation by E-SEM imaging. Based on the bio-nano manipulation system, we propose a nanoinjection method by nanoprobe insertion and nanobead embeddedness. The nanoprobe was designed to have four tips to fix single nanobead. It is fabricated by focused ion beam (FIB) process. Its nano-scale size is considered to be important for minimal damage to a biological specimen. In this work, as biological specimen, the Caenorhabditis elegans (C. elegans) was used. It is one of the important model organisms for various diseases analysis. The nanoinjection technique is applied to transport fluorescent materials or specific biological organism into a specific cell for in-vivo experiment of C. elegans. The proposed system is considered to be important as future nano-surgery system for life innovation using model organism.

This paper presents a novel nano-injection method by nanoprobe insertion based on environmental-scanning electron microscope (E-SEM) under hybrid microscope. The hybrid microscope is designed to combine the optical microscope (OM) and E-SEM to realize biological specimen analysis by optical imaging including fluorescent imaging, and nano-scale manipulation by E-SEM imaging. Based on the bio-nano manipulation system, we propose a nano-injection method by nanoprobe insertion. The nanoprobe is designed to have a sharp tip to insert the probe into biological specimen. It is fabricated by focused ion beam (FIB) process. The nano-scale size is important for minimal damage to biological specimen. The Caenorhabditis elegans (C. elegans) is used as a target biological specimen, which is one of the important model organism for diseases analysis. The nano-injection technique is needed to transport fluorescent materials or specific biological organism into specific cell for in-vivo experiment. The proposed system is considered to be important as future nano-surgery system for life innovation using model organism.