MicroSUCI: A microsurgical background that incorporates suction under continuous irrigation

The microsurgical anastomosis is integral to the success of autologous-free tissue transfer. Successful performance of this procedure relies strongly on operator dexterity, which can be made more challenging when blood and edematous fluids obscure the field of view. Workflow is impeded by intermittent irrigation and suctioning, necessitating presence of an assistant, with risk of arterial thrombosis, from vessels being drawn into suction drains. To negate these current disadvantages and minimize the barrier of entry to microvascular operations, we designed, manufactured, and patented a novel three-dimensional printed microsurgical background device with microfluidic capabilities that allow continuous suction and irrigation as well as provide platforms that enable multiangle retraction to facilitate operator autonomy. This was validated in an ex vivo model, with the device found to be superior to the current standard. We believe that this will have major applicability to the improvement of microsurgeon

Hand-portable HPLC with broadband spectral detection enables analysis of complex polycyclic aromatic hydrocarbon mixtures

Polycyclic aromatic hydrocarbons (PAHs) are considered priority hazardous substances due to their carcinogenic activity and risk to public health. Strict regulations are in place limiting their release into the environment, but enforcement is hampered by a lack of adequate field-testing procedure, instead relying on sending samples to centralised analytical facilities. Reliably monitoring levels of PAHs in the field is a challenge, owing to the lack of field-deployable analytical methods able to separate, identify, and quantify the complex mixtures in which PAHs are typically observed. Here, we report the development of a hand-portable system based on high-performance liquid chromatography incorporating a spectrally wide absorption detector, capable of fingerprinting PAHs based on their characteristic spectral absorption profiles: identifying 100% of the 24 PAHs tested, including full coverage of the United States Environmental Protection Agency priority pollutant list. We report unsupervised methods to exploit these new capabilities for feature detection and identification, robust enough to detect and classify co-eluting and hidden peaks. Identification is fully independent of their characteristic retention times, mitigating matrix effects which can preclude reliable determination of these analytes in challenging samples. We anticipate the platform to enable more sophisticated analytical measurements, supporting real-time decision making in the field

Zero electrical power pump for portable high-performance liquid chromatography

A major trend in analytical chemistry is the miniaturization of laboratory instrumentation. We report a pump requiring no power to operate based on the controlled expansion of a pre-pressurised gas for use in portable applications of high-performance liquid chromatography. The performance of the gas pump is characterised and integrated into a compact liquid chromatography system capable of isocratic separations integrating an LED-based UV-absorption detector. The system weighed 6.7 kg when the mobile phase reservoir was fully charged with 150 mL solvent and included an on-board computer to control the system and analyse data. We characterise the flow-rate through chromatography columns with a variety of geometries and packing materials for a range of pressures up to 150 bar. The maximum variation in flow rate was measured to be 6.5 nL min−1, limited by the resolution of the flow detector. All tests were made on battery power and results are a mixture of those made in the laboratory and in the field. Additionally, we performed a series of 1 m drop tests on the device and show the system's high tolerance to mechanical shocks during operation in the field

3D Printed microvascular clamps: A safe, cheap and effective instrumentation for microsurgery training

Microsurgical training involves practice in ex-vivo models during the early learning curve and poor instrument handling by the inexperienced microsurgeons can cause damage to microsurgical instrumentation or clamps which is particularly costly. To address this, we demonstrate the development, design, manufacturing and application of three different types of 3D printed microvascular clamps in an ex-vivo simulation training model. This report provides evidence of a low cost and easily accessible device that facilitates the process of microsurgical training. The clamps were found to provide advantages akin to normal stainlesssteel microvascular clamps in training setting

Counting proteins in single cells with addressable droplet microarrays

Often cellular behaviour and cellular responses are analysed at the population level where the responses of many cells are pooled together as an average result masking the rich single cell behaviour within a complex population. Single cell protein detection and quantification technologies have made a remarkable impact in recent years. Here we describe a practical and flexible single cell analysis platform based on addressable droplet microarrays. This study describes how the absolute copy numbers of target proteins may be measured with single cell resolution. The tumour suppressor p53 is the most commonly mutated gene in human cancer, with more than 50% of total cancer cases exhibiting a non-healthy p53 expression pattern. The protocol describes steps to create 10nL droplets within which single human cancer cells are isolated and the copy number of p53 protein is measured with single molecule resolution to precisely determine the variability in expression. The method may be applied to any cell type including primary material to determine the absolute copy number of any target proteins of interest

Absolute quantification of protein copy number in single cells with immunofluorescence microscopy calibrated using single molecule microarrays

Great strides toward routine single cell analyses have been made over the last decade, particularly in the field of transcriptomics. For proteomics, amplification is not currently possible and has necessitated the development of ultra-sensitive platforms capable of performing such analyses on single cells. These platforms are improving in terms of throughput and multiplexability but still fall short in relation to more established methods such as fluorescence microscopy. However, microscopy methods rely on fluorescence intensity as a proxy for protein abundance and are not currently capable of reporting this in terms of absolute copy number. Here, a microfluidic implementation of single molecule microarrays for single cell analysis is assessed in its ability to calibrate fluorescence microscopy data. We show that the equivalence of measurements of the steady-state distribution of protein abundance to single molecule microarray data can be exploited to pave the way for absolute quantitation by fluorescence and immunofluorescence microscopy. The methods presented have been developed using GFP but are extendable to other proteins and other biomolecules of interest