The Rorschach butterfly, understanding bone biomechanics prior to using nomenclature in bone trauma interpretations.

Blunt traumas are the most common injuries observed and reported in medical examiner settings. Two common methods to describe bending bone fractures in the anthropological literature include the application of morphology nomenclature and describing characteristic fracture morphology. A nomenclature descriptor of blunt trauma, the butterfly fracture, is commonly used to describe broken long bones. In this paper, a case study of a fractured long bone in a pedestrian vehicle accident is used to highlight the complex interplay of factors involved in bone fracture formation. The application of a butterfly fracture pattern in trauma analysis is useful in establishing the bending direction of a bone, in identifying failure modes, and is valuable in teaching. Yet, butterfly fracture characteristics need to be examined in 3-dimensions for diagnosis of modes of failure, tension, shear and compression, and even then, the bending direction of a broken bone may not provide a reliable indicator of the point of impact (POI); this is especially true when a priori knowledge of the injury is unknown. Common fracture nomenclature, such as oblique, transverse and/or comminuted, as well as eponyms, are medical descriptions of an injury which are impractical to use for interpreting a broken bone from fleshed or skeletonized remains, in establishing a POI and in evaluating total bone trauma (TBT). The examination ofcharacteristic features on the surface of a broken bone associated with the modes of failure is the best approach for establishing the bending direction of a long bone.

Microchip device for performing enzyme assays.

An automated enzyme assay was performed within a microfabricated channel network. Precise concentrations of substrate, enzyme, and inhibitor were mixed in nanoliter volumes using electrokinetic flow. Reagent dilution and mixing were controlled by regulating the applied potential at the terminus of each channel, using voltages derived from an equivalent circuit model of the microchip. The enzyme beta-galactosidase (beta-Gal) was assayed using resorufin beta-D-galactopyranoside (RBG), a substrate that is hydrolyzed to resorufin, a fluorescent product. Reaction kinetics were obtained by varying the concentration of substrate on-chip and monitoring the production of resorufin using laser-induced fluorescence. Derived Michaelis--Menten constants compared well between an on-chip and a conventional enzyme assay. Bias in the derived K(m) and kcat was primarily due to the limited solubility of RBG and the associated lack of measurements at substrate concentrations exceeding the K(m). A Ki of 8 microM for the inhibitor phenylethyl beta-D-thiogalactoside (PETG) was determined from plots of initial rate versus substrate concentration obtained at three concentrations of PETG. The relative inhibition of beta-Gal by lactose, p-hydroxymercuribenzoic acid, and PETG was determined by varying the inhibitor concentration with constant enzyme and substrate concentration. An enzyme assay performed on the microchip within a 20-min period required only 120 pg of enzyme and 7.5 ng of substrate, reducing the amount of reagent consumed by 4 orders of magnitude over a conventional assay.

Electric field directed nucleic acid hybridization on microchips.

Selection and adjustment of proper physical parameters enables rapid DNA transport, site selective concentration, and accelerated hybridization reactions to be carried out on active microelectronic arrays. These physical parameters include DC current, voltage, solution conductivity and buffer species. Generally, at any given current and voltage level, the transport or mobility of DNA is inversely proportional to electrolyte or buffer conductivity. However, only a subset of buffer species produce both rapid transport, site specific concentration and accelerated hybridization. These buffers include zwitterionic and low conductivity species such as: d- and l-histidine; 1- and 3-methylhistidines; carnosine; imidazole; pyridine; and collidine. In contrast, buffers such as glycine, beta-alanine and gamma-amino-butyric acid (GABA) produce rapid transport and site selective concentration but do not facilitate hybridization. Our results suggest that the ability of these buffers (histidine, etc.) to facilitate hybridization appears linked to their ability to provide electric field concentration of DNA; to buffer acidic conditions present at the anode; and in this process acquire a net positive charge which then shields or diminishes repulsion between the DNA strands, thus promoting hybridization.

Continuous separation of high molecular weight compounds using a microliter volume free-flow electrophoresis microstructure.

A microliter volume free-flow electrophoresis microstructure (µ-FFE) was used to perform a continuous separation of high molecular weight compounds. The µ-FFE microstructure had a separation bed volume of 25 µL and was fabricated from silicon using standard micromachining technology. Laser-induced fluorescence was used to detect the sample components, which were labeled with fluorescein isothiocyanate (FITC) prior to analysis. The continuous separation of human serum albumin (HSA), bradykinin, and ribonuclease A demonstrated that only 25 V/cm was required to isolate HSA from bradykinin and ribonuclease A, while 100 V/cm was needed for the separation of bradykinin from ribonuclease A. Comparison of the observed band broadening with the theoretical variance indicated that dispersion due to the initial bandwidth, diffusion, and hydrodynamic broadening were the main contributors to the band broadening of HSA and bradykinin. However, the band broadening for ribonuclease A could not be sufficiently accounted for using the above contributors. Adsorption was found to be a possible contributor to the overall variance for ribonuclease A. Calculation of the theoretical variance due to Joule heating indicated that broadening due to Joule heating effects was insignificant. This was likely due to the narrow cross-sectional area of the device, which facilitated efficient cooling. Electrohydrodynamic distortion was observed for HSA as it migrated toward the side bed. Studies of the resolution of bradykinin and ribonuclease A as a function of field strength at various sample and carrier flow rates indicated that, for maximum throughput, high field strengths and high flow rates were required. However, no optimal conditions were found. The µ-FFE device has a peak capacity of ∼8 bands/cm, while for a typical separation of proteins using a commercial system, a peak capacity of 10 bands/cm is obtained. Thus, the resolving power of the µ-FFE device is similar to those of conventional systems. The continuous separation of tryptic digests of mellitin and cytochrome c demonstrated the ability to continuously separate more complex mixtures. Finally, modifications were made to the microstructure to facilitate fraction collection, and the fractionation of whole rat plasma was performed. Off-line analysis of the resulting fractions indicated that the complete isolation of serum albumin and globulins was possible using a field strength of 25 V/cm.