A scanning electron microscope (SEM) provides real-time imaging with nanometer resolution and a large scanning area, which enables the development and integration of robotic nanomanipulation systems inside a vacuum chamber to realize simultaneous imaging and direct interactions with nanoscaled samples. Emerging techniques for nanorobotic manipulation during SEM imaging enable the characterization of nanomaterials and nanostructures and the prototyping/assembly of nanodevices. This paper presents a comprehensive survey of recent advances in nanorobotic manipulation, including the development of nanomanipulation platforms, tools, changeable toolboxes, sensing units, control strategies, electron beam-induced deposition approaches, automation techniques, and nanomanipulation-enabled applications and discoveries. The limitations of the existing technologies and prospects for new technologies are also discussed.

A long-standing question in natural reproduction is how mammalian sperm navigate inside female reproductive tract and finally reach the egg cell, or oocyte. Recently, fluid flow was proposed as a long–range guidance cue for sperm navigation. Coitus induces fluid flow from oviduct to uterus and sperm align themselves against the flow direction and swim upstream, a phenomenon termed rheotaxis. Whether sperm rheotaxis is a passive process dominated by fluid mechanics, or sperm actively sense and adapt to fluid flow remains controversial. Here we report the first quantitative study of sperm flagellar motion during human sperm rheotaxis and provide direct evidence indicating that sperm rheotaxis is a passive process. Experimental results show that there is no significant difference in flagellar beating amplitude and asymmetry between rheotaxis-turning sperm and those sperm swimming freely in the absence of fluid flow. Additionally, fluorescence image tracking shows no Ca²⁺ influx during sperm rheotaxis turning, further suggesting there is no active signal transduction during human sperm rheotaxis.

Properties of organelles and intracellular structures play important roles in regulating cellular functions, such as gene expression, cell motility and metabolism. The ability to directly interrogate intracellular structures inside a single cell for measurement and manipulation has significant implications in the understanding of subcellular and suborganelle activities, diagnosing diseases, and potentially developing new therapeutic approaches. In the past few decades, a number of technologies have been developed to study single-cell properties. However, methods of measuring intracellular properties and manipulating subcellular structures have been largely underexplored. Due to the even smaller size of intracellular targets and lower signal-to-noise ratio than that in whole-cell studies, the development of tools for intracellular measurement and manipulation is challenging. This paper reviews emerging microsystems and nanoengineered technologies for sensing and quantitative measurement of intracellular properties and for manipulating structures inside a single cell. Recent progress and limitations of these new technologies as well as new discoveries and prospects are discussed.

Compared to robotic injection of suspended cells (e.g., embryos and oocytes), fewer attempts were made to automate the injection of adherent cells (e.g., cancer cells and cardiomyocytes) due to their smaller size, highly irregular morphology, small thickness (a few micrometers thick), and large variations in thickness across cells. This paper presents a robotic system for automated microinjection of adherent cells. The system is embedded with several new capabilities: automatically locating micropipette tips; robustly detecting the contact of micropipette tip with cell culturing surface and directly with cell membrane; and precisely compensating for accumulative positioning errors. These new capabilities make it practical to perform adherent cell microinjection truly via computer mouse clicking in front of a computer monitor, on hundreds and thousands of cells per experiment (versus a few to tens of cells as state of the art). System operation speed, success rate, and cell viability rate were quantitatively evaluated based on robotic microinjection of over 4000 cells. This paper also reports the use of the new robotic system to perform cell-cell communication studies using large sample sizes. The gap junction function in a cardiac muscle cell line (HL-1 cells), for the first time, was quantified with the system.