Automated zebrafish chorion removal and single embryo placement: optimizing throughput of zebrafish developmental toxicity screens

The potential of the developing zebrafish model for toxicology and drug discovery is limited by inefficient approaches to manipulating and chemically exposing zebrafish embryos-namely, manual placement of embryos into 96- or 384-well plates and exposure of embryos while still in the chorion, a barrier of poorly characterized permeability enclosing the developing embryo. We report the automated dechorionation of 1600 embryos at once at 4 h postfertilization (hpf) and placement of the dechorionated embryos into 96-well plates for exposure by 6 hpf. The process removed ≥95% of the embryos from their chorions with 2% embryo mortality by 24 hpf, and 2% of the embryos malformed at 120 hpf. The robotic embryo placement allocated 6-hpf embryos to 94.7% ± 4.2% of the wells in multiple 96-well trials. The rate of embryo mortality was 2.8% (43 of 1536) from robotic handling, the rate of missed wells was 1.2% (18 of 1536), and the frequency of multipicks was <0.1%. Embryo malformations observed at 24 hpf occurred nearly twice as frequently from robotic handling (16 of 864; 1.9%) as from manual pipetting (9 of 864; 1%). There was no statistical difference between the success of performing the embryo placement robotically or manually.     

Real-time evaluation and visualization of learner performance in a mixed-reality environment for clinical breast examination

We investigate the efficacy of incorporating real-time feedback of user performance within mixed-reality environments (MREs) for training real-world tasks with tightly coupled cognitive and psychomotor components. This paper presents an approach to providing real-time evaluation and visual feedback of learner performance in an MRE for training clinical breast examination (CBE). In a user study of experienced and novice CBE practitioners (n = 69), novices receiving real-time feedback performed equivalently or better than more experienced practitioners in the completeness and correctness of the exam. A second user study (n = 8) followed novices through repeated practice of CBE in the MRE. Results indicate that skills improvement in the MRE transfers to the real-world task of CBE of human patients. This initial case study demonstrates the efficacy of MREs incorporating real-time feedback for training real-world cognitive-psychomotor tasks.     

A lysine to arginine substitution at position 146 of rabbit aldolase A changes the rate-determining step to Schiff base formation

Lys146 of rabbit aldolase A [D-fructose-1,6-bis(phosphate):D-glyceraldehyde-3-phosphate lyase, EC 4.1.2.13 [EC] ] was changedto arginine by site-directed mutagenesis. The k_cat of the resultingmutant protein, K146R, was 500 times slower than wild-type insteady-state kinetic assays for both cleavage and condensationof fructose-1,6-bis(phosphate), while the Km for this substratewas unchanged. Analysis of the rate of formation of catalyticintermediates showed K146R was significantly different fromthe wild-type enzyme and other enzymes mutated at this site.Single-turnover experiments using acid precipitation to trapthe Schiff base intermediate on the wild-type enzyme failedto show a build-up of this intermediate on K146R. However, K146Rretained the ability to form the Schiff base intermediate asshown by the significant amounts of Schiff base intermediatetrapped with NaBH₄. In the single-turnover experiments it appearedthat the Schiff base intermediate was converted to productsmore rapidly than it was produced. This suggested a maximalrate of Schiff base formation of 0.022 s^–1, which wasclose to the value of k_cat for this enzyme. This observationis strikingly different from the wild-type enzyme in which Schiffbase formation is >100 times faster than k_cat. For K146Rit appears that steps up to and including Schiff base formationare rate limiting for the catalytic reaction. The carbanionintermediate derived from either substrate or product, and theequilibrium concentrations of covalent enzyme-substrate intermediates,were much lower on K146R than on the wild-type enzyme. The greaterbulk of the guanidino moiety may destabilize the covalent enzyme-substrateintermediates, thereby slowing the rate of Schiff base formationsuch that it becomes rate limiting. The K146R mutant enzymeis significantly more active than other enzymes mutated at thissite, perhaps because it maintains a positively charged groupat an essential position in the active site or perhaps the Argfunctionally substitutes as a general acid/base catalyst inboth Schiff base formation and in subsequent abstraction ofthe C4-hydroxyl proton.     

The Coolest Way to Generate Binary Strings

Pick a binary string of length n and remove its first bit b. Now insert b after the first remaining 10, or insert $\overline{b}$ at the end if there is no remaining 10. Do it again. And again. Keep going! Eventually, you will cycle through all 2 ⁿ of the binary strings of length n. For example, are the binary strings of length n=4, where and . And if you only want strings with weight (number of 1s) between ? and u? Just insert b instead of $\overline{b}$ when the result would have too many 1s or too few 1s. For example, are the strings with n=4, ?=0 and u=2. This generalizes ‘cool-lex’ order by Ruskey and Williams (The coolest way to generate combinations, Discrete Mathematics) and we present two applications of our ‘cooler’ order. First, we give a loopless algorithm for generating binary strings with any weight range in which successive strings have Levenshtein distance two. Second, we construct de Bruijn sequences for (i) ?=0 and any u (maximum specified weight), (ii) any ? and u=n (minimum specified weight), and (iii) odd u?? (even size weight range). For example, all binary strings with n=6, ?=1, and u=4 appear once (cyclically) in . We also investigate the recursive structure of our order and show that it shares certain sublist properties with lexicographic order.     

Rapid consolidation of powdered materials by induction hot pressing

A rapid hot press system in which the heat is supplied by RF induction to rapidly consolidate thermoelectric materials is described. Use of RF induction heating enables rapid heating and consolidation of powdered materials over a wide temperature range. Such rapid consolidation in nanomaterials is typically performed by spark plasma sintering (SPS) which can be much more expensive. Details of the system design, instrumentation, and performance using a thermoelectric material as an example are reported. The Seebeck coefficient, electrical resistivity, and thermal diffusivity of thermoelectric PbTe material pressed at an optimized temperature and time in this system are shown to agree with material consolidated under typical consolidation parameters.     

A high temperature apparatus for measurement of the Seebeck coefficient

A high temperature Seebeck coefficient measurement apparatus with various features to minimize typical sources of error is designed and built. Common sources of temperature and voltage measurement error are described and principles to overcome these are proposed. With these guiding principles, a high temperature Seebeck measurement apparatus with a uniaxial 4-point contact geometry is designed to operate from room temperature to over 1200 K. This instrument design is simple to operate, and suitable for bulk samples with a broad range of physical types and shapes.     

An Empirical Evaluation of the System Usability Scale

This article presents nearly 10 year's worth of System Usability Scale (SUS) data collected on numerous products in all phases of the development lifecycle. The SUS, developed by Brooke (1996) Brooke, J. 1996. “SUS: A “quick and dirty” usability scale”. In Usability evaluation in industry, Edited by: Jordan, P. W., Thomas, B. A. Weerdmeester and McClelland, I. L. 189–194. London: Taylor & Francis.  [Google Scholar], reflected a strong need in the usability community for a tool that could quickly and easily collect a user's subjective rating of a product's usability. The data in this study indicate that the SUS fulfills that need. Results from the analysis of this large number of SUS scores show that the SUS is a highly robust and versatile tool for usability professionals. The article presents these results and discusses their implications, describes nontraditional uses of the SUS, explains a proposed modification to the SUS to provide an adjective rating that correlates with a given score, and provides details of what constitutes an acceptable SUS score.     

Nonrigid structure-from-motion: estimating shape and motion with hierarchical Priors

This paper describes methods for recovering time-varying shape and motion of non-rigid 3D objects from uncalibrated 2D point tracks. For example, given a video recording of a talking person, we would like to estimate the 3D shape of the face at each instant, and learn a model of facial deformation. Time-varying shape is modeled as a rigid transformation combined with a non-rigid deformation. Reconstruction is ill-posed if arbitrary deformations are allowed, and thus additional assumptions about deformations are required. We first suggest restricting shapes to lie within a low-dimensional subspace, and describe estimation algorithms. However, this restriction alone is insufficient to constrain reconstruction. To address these problems, we propose a reconstruction method using a Probabilistic Principal Components Analysis (PPCA) shape model, and an estimation algorithm that simultaneously estimates 3D shape and motion for each instant, learns the PPCA model parameters, and robustly fills-in missing data points. We then extend the model to model temporal dynamics in object shape, allowing the algorithm to robustly handle severe cases of missing data.