Learning and Transfer in Problem Solving Progressions.

Do individuals learn more effectively when given progressive or variable problem-solving experience, relative to consistent problem-solving experience? We investigated this question using a Rubik's Cube paradigm. Participants were randomly assigned to a progression-order condition, where they practiced solving three progressively more difficult Rubik's Cubes (i.e., 2 × 2 × 2 to 3 × 3 × 3 to 4 × 4 × 4), a variable-order condition, where they practiced solving three Rubik's Cubes of varying difficulty (e.g., 3 × 3 × 3 to 2 × 2 × 2 to 4 × 4 × 4), or a consistent-order condition, where they consistently practiced on three 5 × 5 × 5 Rubik's Cubes. All the participants then attempted a 5 × 5 × 5 Rubik's Cube test. We tested whether variable training is as effective as progressive training for near transfer of spatial skills and whether progressive training is superior to consistent training. We found no significant differences in performance across conditions. Participants' fluid reasoning predicted 5 × 5 × 5 Rubik's Cube test performance regardless of training condition.

Multiview confocal super-resolution microscopy.

Confocal microscopy1 remains a major workhorse in biomedical optical microscopy owing to its reliability and flexibility in imaging various samples, but suffers from substantial point spread function anisotropy, diffraction-limited resolution, depth-dependent degradation in scattering samples and volumetric bleaching2. Here we address these problems, enhancing confocal microscopy performance from the sub-micrometre to millimetre spatial scale and the millisecond to hour temporal scale, improving both lateral and axial resolution more than twofold while simultaneously reducing phototoxicity. We achieve these gains using an integrated, four-pronged approach: (1) developing compact line scanners that enable sensitive, rapid, diffraction-limited imaging over large areas; (2) combining line-scanning with multiview imaging, developing reconstruction algorithms that improve resolution isotropy and recover signal otherwise lost to scattering; (3) adapting techniques from structured illumination microscopy, achieving super-resolution imaging in densely labelled, thick samples; (4) synergizing deep learning with these advances, further improving imaging speed, resolution and duration. We demonstrate these capabilities on more than 20 distinct fixed and live samples, including protein distributions in single cells; nuclei and developing neurons in Caenorhabditis elegans embryos, larvae and adults; myoblasts in imaginal disks of Drosophila wings; and mouse renal, oesophageal, cardiac and brain tissues.

Multi-immersion open-top light-sheet microscope for high-throughput imaging of cleared tissues.

Recent advances in optical clearing and light-sheet microscopy have provided unprecedented access to structural and molecular information from intact tissues. However, current light-sheet microscopes have imposed constraints on the size, shape, number of specimens, and compatibility with various clearing protocols. Here we present a multi-immersion open-top light-sheet microscope that enables simple mounting of multiple specimens processed with a variety of clearing protocols, which will facilitate wide adoption by preclinical researchers and clinical laboratories. In particular, the open-top geometry provides unsurpassed versatility to interface with a wide range of accessory technologies in the future.

Scalable Nanogap Sensors for Non-Redox Enzyme Assays.

Clinical diagnostic assays that monitor redox enzyme activity are widely used in small, low-cost readout devices for point-of-care monitoring (e.g., a glucometer); however, monitoring non-redox enzymes in real-time using compact electronic devices remains a challenge. We address this problem by using a highly scalable nanogap sensor array to observe electrochemical signals generated by a model non-redox enzyme system, the DNA polymerase-catalyzed incorporation of four modified, redox-tagged nucleotides. Using deoxynucleoside triphosphates (dNTPs) tagged with para-aminophenyl monophosphate (pAPP) to form pAP-deoxyribonucleoside tetra-phosphates (AP-dN4Ps), incorporation of the nucleotide analogs by DNA polymerase results in the release of redox inactive pAP-triphosphates (pAPP3) that are converted to redox active small molecules para-aminophenol (pAP) in the presence of phosphatase. In this work, cyclic enzymatic reactions that generated many copies of pAP at each base incorporation site of a DNA template in combination with the highly confined nature of the planar nanogap transducers ( z = 50 nm) produced electrochemical signals that were amplified up to 100,000×. We observed that the maximum signal level and amplification level were dependent on a combination of factors including the base structure of the incorporated nucleotide analogs, nanogap electrode materials, and electrode surface coating. In addition, electrochemical signal amplification by redox cycling in the nanogap is independent of the in-plane geometry of the transducer, thus allowing the nanogap sensors to be highly scalable. Finally, when the DNA template concentration was constrained, the DNA polymerase assay exhibited different zero-order reaction kinetics for each type of base incorporation reaction, resolving the closely related nucleotide analogs.

Visualizing Calcium Flux in Freely Moving Nematode Embryos.

The lack of physiological recordings from Caenorhabditis elegans embryos stands in stark contrast to the comprehensive anatomical and gene expression datasets already available. Using light-sheet fluorescence microscopy to address the challenges associated with functional imaging at this developmental stage, we recorded calcium dynamics in muscles and neurons and developed analysis strategies to relate activity and movement. In muscles, we found that the initiation of twitching was associated with a spreading calcium wave in a dorsal muscle bundle. Correlated activity in muscle bundles was linked with early twitching and eventual coordinated movement. To identify neuronal correlates of behavior, we monitored brainwide activity with subcellular resolution and identified a particularly active cell associated with muscle contractions. Finally, imaging neurons of a well-defined adult motor circuit, we found that reversals in the eggshell correlated with calcium transients in AVA interneurons.

Label-free electrical detection of pyrophosphate generated from DNA polymerase reactions on field-effect devices.

We introduce a label-free approach for sensing polymerase reactions on deoxyribonucleic acid (DNA) using a chelator-modified silicon-on-insulator field-effect transistor (SOI-FET) that exhibits selective and reversible electrical response to pyrophosphate anions. The chemical modification of the sensor surface was designed to include rolling-circle amplification (RCA) DNA colonies for locally enhanced pyrophosphate (PPi) signal generation and sensors with immobilized chelators for capture and surface-sensitive detection of diffusible reaction by-products. While detecting arrays of enzymatic base incorporation reactions is typically accomplished using optical fluorescence or chemiluminescence techniques, our results suggest that it is possible to develop scalable and portable PPi-specific sensors and platforms for broad biomedical applications such as DNA sequencing and microbe detection using surface-sensitive electrical readout techniques.

High-k dielectric Al2O3 nanowire and nanoplate field effect sensors for improved pH sensing.

Over the last decade, field-effect transistors (FETs) with nanoscale dimensions have emerged as possible label-free biological and chemical sensors capable of highly sensitive detection of various entities and processes. While significant progress has been made towards improving their sensitivity, much is yet to be explored in the study of various critical parameters, such as the choice of a sensing dielectric, the choice of applied front and back gate biases, the design of the device dimensions, and many others. In this work, we present a process to fabricate nanowire and nanoplate FETs with Al(2)O(3) gate dielectrics and we compare these devices with FETs with SiO(2) gate dielectrics. The use of a high-k dielectric such as Al(2)O(3) allows for the physical thickness of the gate dielectric to be thicker without losing sensitivity to charge, which then reduces leakage currents and results in devices that are highly robust in fluid. This optimized process results in devices stable for up to 8 h in fluidic environments. Using pH sensing as a benchmark, we show the importance of optimizing the device bias, particularly the back gate bias which modulates the effective channel thickness. We also demonstrate that devices with Al(2)O(3) gate dielectrics exhibit superior sensitivity to pH when compared to devices with SiO(2) gate dielectrics. Finally, we show that when the effective electrical silicon channel thickness is on the order of the Debye length, device response to pH is virtually independent of device width. These silicon FET sensors could become integral components of future silicon based Lab on Chip systems.

Simultaneous measurement of nonlinearity and electrochemical impedance for protein sensing using two-tone excitation.

Impedance biosensors detect the binding of a target to an immobilized probe by quantifying changes in the impedance of the electrode-electrolyte interface. The interface's I-V relationship is inherently nonlinear, varying with DC bias, and target binding can alter the degree of nonlinearity. We propose and demonstrate a method to simultaneously measure the nonlinearity and conventional small-signal impedance using intermodulation products from a two-tone input. Intermodulation amplitudes accurately reflect the impedance's manually-measured voltage dependence. We demonstrate that changes in nonlinearity can discriminate protein binding. Our measurements suggest that target binding can alter nonlinearity via the voltage dependence of the ionic double layer.

A System for Multiplexed Direct Electrical Detection of DNA Synthesis.

An electronic system for the multiplexed detection of DNA polymerization is designed and characterized. DNA polymerization is detected by the measurement of small transient currents arising from ion diffusion during polymerization. A transimpedance amplifier is used to detect these small currents; we implemented a twenty-four channel recording system on a single printed circuit board. Various contributions to the input-referred current noise are analyzed and characterized, as it limits the minimum detectable current and thus the biological limit of detection. We obtained 8.5 pA RMS mean noise current (averaged over all 24 channels) over the recording bandwidth (DC to 2 kHz). With digital filtering, the input-referred current noise of the acquisition system is reduced to 2.4 pA, which is much lower than the biological noise. Electrical crosstalk between channels is measured, and a model for the crosstalk is presented. Minimizing the crosstalk is critical because it can lead to erroneous microarray data. With proper precautions, crosstalk is reduced to a negligible value (less than 1.4%). Using a micro-fabricated array of 24 gold electrodes, we demonstrated system functionality by detecting the presence of a target DNA oligonucleotide which hybridized onto its corresponding target.

Label-Free Impedance Biosensors: Opportunities and Challenges.

Impedance biosensors are a class of electrical biosensors that show promise for point-of-care and other applications due to low cost, ease of miniaturization, and label-free operation. Unlabeled DNA and protein targets can be detected by monitoring changes in surface impedance when a target molecule binds to an immobilized probe. The affinity capture step leads to challenges shared by all label-free affinity biosensors; these challenges are discussed along with others unique to impedance readout. Various possible mechanisms for impedance change upon target binding are discussed. We critically summarize accomplishments of past label-free impedance biosensors and identify areas for future research.