As a fundamental property of light, the angular momentum of photons has been of great interest. Here, we demonstrate that optical spin-to-orbital angular momentum conversion can occur in a homogeneous and isotropic medium. This Letter presents both theoretical and experimental studies of this conversion in a tightly focused beam and shows that the orbital rotation speeds of trapped particles are altered because of this conversion as predicted by theory.

This paper describes a method, which combines optical trapping and microfluidic-based droplet generation, for selectively and controllably encapsulating a single target cell or subcellular structure, such as a mitochondrion, into a picoliter- or femtoliter-volume aqueous droplet that is surrounded by an immiscible phase. Once the selected cell or organelle is encased within the droplet, it is stably confined in the droplet and cannot be removed. We demonstrate in droplet the rapid laser photolysis of the single cell, which essentially "freezes" the state that the cell was in at the moment of photolysis and confines the lysate within the small volume of the droplet. Using fluorescein di-beta-d-galactopyranoside, which is a fluorogenic substrate for the intracellular enzyme beta-galactosidase, we also assayed the activity of this enzyme from a single cell following the laser-induced lysis of the cell. This ability to entrap individual selected cells or subcellular organelles should open new possibilities for carrying out single-cell studies and single-organelle measurements.

By using methods that permit the generation and manipulation of ultrasmall-volume droplets, researchers are pushing the boundaries of ultrasensitive chemical analyses. (To listen to a podcast about this feature, please go to the Analytical Chemistry Web site at pubs.acs.org/ancham.)

Intraneuronal accumulation of amyloid-β (Aβ) peptides represent an early pathological feature in Alzheimer’s disease. We have therefore utilized flow cytometry and confocal microscopy in combination with endocytosis inhibition to explore the internalisation efficiency and uptake mechanisms of Aβ(1–40) and Aβ(1–42) monomers in cultured SH-SY5Y cells. We find that both variants are constitutively internalised via endocytosis and that their uptake is proportional to cellular endocytic rate. Moreover, SH-SY5Y cells internalise consistently twice the amount of Aβ(1–42) compared to Aβ(1–40); an imaging-based quantification showed that cells treated with 1 µM peptide for 8 h contained 800,000 peptides of Aβ(1–42) and 400,000 of Aβ(1–40). Both variants co-localised to >90% with lysosomes or other acidic compartments. Dynasore and chlorpromazine endocytosis inhibitors were both found to reduce uptake, particularly of Aβ(1–42). Overexpression of the C-terminal of the clathrin-binding domain of AP180, dynamin2 K44A, or Arf6 Q67L did however not reduce uptake of the Aβ variants. By contrast, perturbation of actin polymerisation and inhibition of macropinocytosis reduced Aβ(1–40) and Aβ(1–42) uptake considerably. This study clarifies mechanisms of Aβ(1–40) and Aβ(1–42) uptake, pinpoints differences between the two variants and highlights a common and putative role of macropinocytosis in the early accumulation of intraneuronal Aβ in AD.

Microfluidics has emerged as a powerful laboratory toolbox for biologists, allowing manipulation and analysis of processes at a cellular and sub-cellular level, through utilization of microfabricated features at size-scales relevant to that of a single cell. In the majority of microfluidic devices, sample processing and analysis occur within closed microchannels, imposing restrictions on sample preparation and use. We present an optimized non-contact open-volume microfluidic tool to overcome these and other restrictions, through the use of a hydrodynamically confined microflow pipette, serving as a multifunctional solution handling and dispensing tool. The geometries of the tool have been optimised for use in optical microscopy, with integrated solution reservoirs to reduce reagent use, contamination risks and cleaning requirements. Device performance was characterised using both epifluorescence and total internal reflection fluorescence (TIRF) microscopy, resulting in ~200 ms and ~130 ms exchange times at ~100 nm and ~30 µm distances to the surface respectively.

This paper describes a fluidic and optical platform for the generation and manipulation of single femtoliter-volume aqueous droplets. Individual droplets were generated on-demand using a microfluidic chamber that confers environmental flow stability. Optical vortex traps were implemented to manipulate and transport the generated droplets, which have a lower refractive index than the immiscible medium in which the droplets are immersed and thus cannot be trapped using conventional optical tweezers. We also demonstrated the ability to shrink and increase the refractive index of the generated droplet, thereby permitting its facile fusion with another droplet using an optical tweezer. To illustrate the versatility of this platform, we have performed both fast (<1 s) and slow (>1 h) chemical reactions in these femtoliter-volume aqueous droplets.

Single-cell nanosurgery and the ability to manipulate nanometer-sized subcellular structures with optical tweezers has widespread applications in biology, but so far has been limited by difficulties in maintaining the functionality of the transported subcellular organelles. This difficulty arises because of the propensity of optical tweezers to photodamage the trapped object. To address this issue, this paper describes the use of a polarization-shaped optical vortex trap, which exerts less photodamage on the trapped particle than conventional optical tweezers, for carrying out single-cell nanosurgical procedures. This method is also anticipated to find broad use in the trapping of any nanoparticles that are adversely affected by high-intensity laser light.Despite the small size of a mammalian cell, it is an extremely heterogeneous and compartmentalized structure. Proteins and small-molecule metabolites constantly traffic among these intracellular compartments, and it has become increasingly evident that biological specificity (e.g. between proteins) relies heavily on spatial and temporal segregation and compartmentalization of molecules in addition to chemical and molecular specificity 1, 2 . Gaining information with regard to the spatial and temporal distribution and evolution of molecules within cells, therefore, is crucial to the construction of a quantitative model of cellular function. The ability to isolate selectively single subcellular compartments for chemical analysis or transplantation opens new venues for studying the spatial and temporal organization of the cell. For example, the reprogramming of the nucleus of somatic cells may be achieved via nuclear transfer 3, 4 . This paper describes and compares the use of polarizationshaped vortex traps with a Gaussian optical tweezer for performing single-cell nanosurgery.Single-beam optical gradient traps, or optical tweezers, have made significant impact on biophysical and biological research in the past two decades 5-11 . Unfortunately, while optical tweezers offer exquisite sensitivity in its ability to position microparticles and to measure the forces exerted by biological motors 12 , it suffers from one important disadvantage: the trapped particle is localized at the laser focus where light intensity is the highest, often reaching 10 7 -10 8 W/cm 2 . As a result, the laser light used to trap a particle also has a propensity to photobleach and photodamage the particle, especially when the particle is fragile and small (e.g. a subcellular organelle that is fluorescently labeled) for which high laser intensities are often required. To minimize radiation damage to the trapped biological particle, laser wavelengths between ∼800nm to ∼ 1100nm are usually used because of the low absorption cross section of water and biological molecules in this spectral range 6-8, 13-15 . Nevertheless, at the high laser intensities required for trapping and translating subcellular organelles through the dense * Corresponding author. E-mail: chiu@chem.washington.edu. ...

This paper describes the use of an optical vortex trap for the transport and fusion of single femtolitervolume aqueous droplets. Individual droplets were generated by emulsifying water in acetophenone with SPAN 80 surfactant. We demonstrate the ability of optical vortex traps to position trapped droplets precisely while excluding surrounding aqueous droplets from entering the trap, thereby preventing unwanted cross contamination by other nearby droplets. Additionally, the limitation of optical vortex traps for inducing droplet fusion is illustrated, and a remedy is provided through modulation of the spatial intensity profile of the optical vortex beam. Spatial modulation was achieved by translating the computer generated hologram (CGH) with respect to the input Gaussian beam, thereby shifting the location of the embedded phase singularity (dark core) within the optical vortex beam. We present both simulated and experimentally measured intensity profiles of the vortex beam caused by translation of the CGH. We further describe the use of this technique to achieve controlled and facile fusion of two aqueous droplets.