We present an imaging technique using an optically trapped cigar-shaped probe controlled using holographic optical tweezers. The probe is raster scanned over a surface, allowing an image to be taken in a manner analogous to scanning probe microscopy (SPM), with automatic closed loop feedback control provided by analysis of the probe position recorded using a high speed CMOS camera. The probe is held using two optical traps centred at least 10 µm from the ends, minimizing laser illumination of the tip, so reducing the chance of optical damage to delicate samples. The technique imparts less force on samples than contact SPM techniques, and allows highly curved and strongly scattering samples to be imaged, which present difficulties for imaging using photonic force microscopy. To calibrate our technique, we first image a known sample--the interface between two 8 µm polystyrene beads. We then demonstrate the advantages of this technique by imaging the surface of the soft alga Pseudopediastrum. The scattering force of our laser applied directly onto this sample is enough to remove it from the surface, but we can use our technique to image the algal surface with minimal disruption while it is alive, not adhered and in physiological conditions. The resolution is currently equivalent to confocal microscopy, but as our technique is not diffraction limited, there is scope for significant improvement by reducing the tip diameter and limiting the thermal motion of the probe.

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PACS 87.80.Cc-Optical trapping PACS 81.07.-b-Nanoscale materials and structures: fabrication and characterization PACS 45.40.-f-Dynamics and kinematics of rigid bodies Abstract-We analyse the thermal motion of a holographically trapped non-spherical force probe, capable of interrogating arbitrary samples with nanometer resolution. High speed video stereo-microscopy is used to track the translational and rotational coordinates of the micro-tool in three dimensions, and the complete 6 × 6 stiffness matrix for the system is determined using equipartition theorem. The Brownian motion of the extended structure is described in terms of a continuous distribution of thermal ellipsoids. A centre of optical stress, at which rotational and translational motion is uncoupled, is observed and controlled. Once calibrated, the micro-tool is deployed in two modes of operation: as a force sensor with < 150 femto-Newton sensitivity, and in a novel form of photonic force microscopy.

Optical trapping techniques have been used to investigate fundamental biological processes ranging from the identification of the processive mechanisms of kinesin and myosin to understanding the mechanics of DNA. To date, these investigations have relied almost exclusively on the use of isotropic probes based on colloidal microspheres. However, there are many potential advantages in utilizing more complex probe morphologies: use of multiple trapping points enables control of the interaction volume; increasing the distance between the optical trap and the sample minimizes photodamage in sensitive biological materials; and geometric anisotropy introduces the potential for asymmetric surface chemistry and multifunctional probes. Here we demonstrate that living cells of the freshwater diatom Nitzschia subacicularis Hustedt can be exploited as advanced probes for holographic optical tweezing applications. We characterize the optical and material properties associated with the high shape anisotropy of the silica frustule, examine the trapping behavior of the living algal cells, and demonstrate how the diatoms can be calibrated for use as force sensors and as force probes in the presence of rat B-cell hybridoma (11B11) cells.

Holographically trapped nanotools can be used in a novel form of force microscopy. By measuring the displacement of the tool in the optical traps, the contact force experienced by the probe can be inferred. In the following paper we experimentally demonstrate the calibration of such a device and show that its behaviour is independent of small changes in the relative position of the optical traps. Furthermore, we explore more general aspects of the thermal motion of the tool.