Water droplet-in-oil digestion method for single-cell proteomics

Recent advances in single-cell proteomics highlight the promise of sensitive analyses in limited cell populations. However, technical challenges remain for sample recovery, throughput, and versatility. Here, we first report a water droplet-in-oil digestion (WinO) method based on carboxyl-coated beads and phase transfer surfactants for proteomic analysis using limited sample amounts. This method was developed to minimize the contact area between the sample solution and the container to reduce the loss of proteins and peptides by adsorption. This method increased protein and peptide recovery 10-fold as well as the number of quantified transmembrane proteins compared to an in-solution digestion (ISD) method. The proteome profiles obtained from 100 cells using the WinO method highly correlated with those from 10000 cells using the ISD method. We successfully applied the WinO method to single-cell proteomics and quantified 462 proteins. Using the WinO method, samples can be easily prepared in a multi-well plate, making it a widely applicable and suitable method for single-cell proteomics.

A large-volume sputum dry storage and transportation device for molecular and culture-based diagnosis of tuberculosis

Technologies for preservation of specimens in the absence of cold chains are essential for optimum utilization of existing laboratory services in the developing world. We present a prototype called specimen transportation tube (SPECTRA-tube) for the collection, exposure-free drying, ambient transportation, and liquid state recovery of large-volume (>1 mL) specimens. Specimens introduced into SPECTRA-tube are dried in glass fiber membranes, which are critical for efficient liquid-state sample recovery by rehydration and centrifugation. Mycobacterium smegmatis (Msm)-spiked mock sputum dried in native Standard 17 glass fiber was stable for molecular testing after 10-day storage at 45°C, and for culture testing after 10- and 5-day storage at 37°C and 45°C, respectively. Compatibility with human sputum storage was demonstrated by dry storing Mycobacterium bovis-spiked pooled human sputum in SPECTRA-tube for 5 days at room temperature followed by successful qPCR detection. By significantly increasing the volume of samples that can be transported in the dry state and enabling recovery of the entire sample in liquid state, SPECTRA-tube presents a potential universal solution for the preservation and transportation of liquid specimens.

Overparameterized neural networks implement associative memory

Identifying computational mechanisms for memorization and retrieval of data is a long-standing problem at the intersection of machine learning and neuroscience. Our main finding is that standard overparameterized deep neural networks trained using standard optimization methods implement such a mechanism for real-valued data. We provide empirical evidence that 1) overparameterized autoencoders store training samples as attractors and thus iterating the learned map leads to sample recovery, and that 2) the same mechanism allows for encoding sequences of examples and serves as an even more efficient mechanism for memory than autoencoding. Theoretically, we prove that when trained on a single example, autoencoders store the example as an attractor. Lastly, by treating a sequence encoder as a composition of maps, we prove that sequence encoding provides a more efficient mechanism for memory than autoencoding.

Operative and Technical Modifications to the Coriolis® µ Air Sampler That Improve Sample Recovery and Biosafety During Microbiological Air Sampling

Detecting infectious aerosols is central for gauging and countering airborne threats. In this regard, the Coriolis® µ cyclonic air sampler is a practical, commercial collector that can be used with various analysis methods to monitor pathogens in air. However, information on how to operate this unit under optimal sampling and biosafety conditions is limited. We investigated Coriolis performance in aerosol dispersal experiments with polystyrene microspheres and Bacillus globigii spores. We report inconsistent sample recovery from the collector cone due to loss of material when sampling continuously for more than 30 min. Introducing a new collector cone every 10 min improved this shortcoming. Moreover, we found that several surfaces on the device become contaminated during sampling. Adapting a high efficiency particulate air-filter system to the Coriolis prevented contamination without altering collection efficiency or tactical deployment. A Coriolis modified with these operative and technical improvements was used to collect aerosols carrying microspheres released inside a Biosafety Level-3 laboratory during simulations of microbiological spills and aerosol dispersals. In summary, we provide operative and technical solutions to the Coriolis that optimize microbiological air sampling and improve biosafety.

Operative and technical modifications to the Coriolis® µ air sampler that improve sample recovery and biosafety during microbiological air sampling

ABSTRACTDetecting infectious aerosols is central for gauging and countering airborne threats. In this regard the Coriolis® µ cyclonic air sampler is a practical, commercial collector that can be used with various analysis methods to monitor pathogens in air. However, information on how to operate this unit under optimal sampling and biosafety conditions is limited. We investigated Coriolis performance in aerosol dispersal experiments with polystyrene microspheres and Bacillus globigii spores. We report inconsistent sample recovery from the collector cone due to loss of material when sampling continuously for more than 30 min. Introducing a new collector cone every 10 min improved this shortcoming. Moreover, we found that several surfaces on the device become contaminated during sampling. Adapting a HEPA-filter system to the Coriolis prevented contamination without altering collection efficiency or tactical deployment. A Coriolis modified with these operative and technical improvements was used to collect aerosols carrying microspheres released inside a Biosafety Level-3 laboratory during simulations of microbiological spills and aerosol dispersals. In summary, we provide operative and technical solutions to the Coriolis that optimize microbiological air sampling and improve biosafety.

Simultaneous Determination of Contents of Flavonol Glycosides and Terpene Lactones in Ginkgo Biloba Tablets by Ultra High Performance Liquid Chromatography Tandem Single Quadrupole Mass Spectrometry Detector

Ginkgo biloba leaf tablets is an effective ingredient in the treatment of cardiovascular and cerebrovascular diseases. In the process of drug production, the quality of ginkgo preparations is often controlled by measuring the content of seven ingredients in ginkgo leaves. To establish UPLC-MS multicomponent analysis method for ginkgo biloba tablets and to simultaneously determine the contents of quercetin (QUE), isorhamnetin(ISO), kaempferol(KAE) and GinkgolideA (GA),ginkgolideB(GB),ginkgolideC(GC) and bilobalide (BB) in ginkgo tablets. Waters Xbridge C18(4.6×150mm,3.5um) column was used, mobile phase A was acetonitrile and mobile phase B was water (containing 0.10% formic acid). The injection volume was 10μL.Negative ion mode monitoring was conducted with ESI. Scanning range:m/z100∼1400.The detection ions of the seven tested components includem/z301.0(QUE),m/z284.9(KAE),m/z315.1(ISO),m/z453.1(GA),m/z423.1(G B),m/z439.0(GC)and m/z325.0(BB), respectively. Within a space of 10min, flavonoids and terpene lactones in ginkgo biloba tablets were completely separated. The peak area exhibited an excellent linear relationship with the concentration. The sample recovery rate ranged from 91.74% to 109.77%.Precision RSDs of within-day and between-day were lower than 2.879% and 3.928% respectively. The method for determination of seven components in ginkgo biloba tablets displays good repeatability, recovery rate and precision, for which it can be applied to quality control of ginkgo biloba tablets.

Droplets for Sampling and Transport of Chemical Signals in Biosensing: A Review

The chemical, temporal, and spatial resolution of chemical signals that are sampled and transported with continuous flow is limited because of Taylor dispersion. Droplets have been used to solve this problem by digitizing chemical signals into discrete segments that can be transported for a long distance or a long time without loss of chemical, temporal or spatial precision. In this review, we describe Taylor dispersion, sampling theory, and Laplace pressure, and give examples of sampling probes that have used droplets to sample or/and transport fluid from a continuous medium, such as cell culture or nerve tissue, for external analysis. The examples are categorized, as follows: (1) Aqueous-phase sampling with downstream droplet formation; (2) preformed droplets for sampling; and (3) droplets formed near the analyte source. Finally, strategies for downstream sample recovery for conventional analysis are described.

Simultaneous Determination of Thirteen Q-Markers in Raw and Processed Tussilago farfara L. by UPLC-QQQ-MS/MS Coupled with Chemometrics

The purpose of this study was to establish a rapid, reliable, and sensitive ultra-performance liquid chromatography with triple-quadrupole tandem mass spectrometry coupled with chemometric method to measure and evaluate the differences between thirteen compounds in raw and processed Tussilago farfara L. from different sources. This assay method was validated, and the results indicated that the calibration curves for the thirteen compounds had good linearity (R2 > 0.9990). The limits of detection and limits of quantification of the thirteen compounds ranged from 0.0012 to 0.0095 μg/mL and from 0.0038 to 0.0316 μg/mL, respectively. The relative standard deviations (RSD) of the intra- and inter-day precisions and stability ranged from 1.06 to 2.00%, 0.26 to 1.99%, and 0.75 to 1.97%, respectively. The sample recovery rates of the thirteen compounds with different concentrations were 94.47–104.06%. The chemometric results, including principal component analysis, hierarchical clustering analysis, three-dimensional analysis, and box plot analysis, indicated that there are significance differences in raw and processed Tussilago farfara L. The results of this study confirm that the proposed method is the first reported method that has been successfully applied for simultaneous determination and discovery of the difference between thirteen compounds of raw and processed Tussilago farfara L. Thus, this method could be a helpful tool for the detection and confirmation of the quality of traditional Chinese medicines and provide a basis for future pharmacological studies.

Determination of permeability of overconsolidated clay from pressuremeter pressure hold tests

A method was developed to interpret the horizontal permeability (kh) from pressuremeter pressure hold tests (PHTs) of approximately 3 min duration. The method relies on a regression analysis of the numerical analysis simulating the consolidation of clay under a constant pressure boundary during undrained expansion. The method was applied to a series of PHTs performed in deep clay formations in the Seattle area. The interpreted permeabilities are thought to be more representative of in situ conditions than those determined by laboratory testing by virtue of reduced disturbance during sample recovery and preparation. Results could be improved with a further exclusion of the creep effect on PHTs.