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HER2 is a crucial therapeutic target in breast cancer, and the survival rate of breast cancer patients has increased because of this receptor's inhibition. However, tumors have shown resistance to this therapeutic strategy due to oncogenic mutations that decrease the binding of several HER2-targeted drugs, including lapatinib, and confer resistance to this drug. Neratinib can overcome this drug resistance and effectively inhibit HER2 signaling and tumor growth. In the present study, we examined the efficacy of lapatinib and neratinib using breast cancer cells by Raman microscopy combined with a deep wavelet scattering-based multivariate analysis framework. This approach discriminated between control cells and drug-treated cells with high accuracy, compared to classical principal component analysis. Both lapatinib and neratinib induced changes in the cellular biochemical composition. Furthermore, the Raman results were compared with the results of several in vitro assays. For instance, drug-treated cells exhibited (i) inhibition of ERK and AKT phosphorylation, (ii) inhibition of cellular proliferation, (iii) cell-cycle arrest, and (iv) apoptosis as indicated by western blotting, real-time cell analysis (RTCA), cell-cycle analysis, and apoptosis assays. Thus, the observed Raman spectral changes are attributed to cell-cycle arrest and apoptosis. The results also indicated that neratinib is more potent than lapatinib. Moreover, the uptake and distribution of lapatinib in cells were visualized through its label-free marker bands in the fingerprint region using Raman spectral imaging. These results show the prospects of Raman microscopy in drug evaluation and presumably in drug discovery.

Linked-read sequencing promises a one-method approach for genome-wide insights including single nucleotide variants (SNVs), structural variants, and haplotyping. We introduce Barcode Linked Reads (BLR), an open-source haplotyping pipeline capable of handling millions of barcodes and data from multiple linked-read technologies including DBS, 10× Genomics, TELL-seq and stLFR. Running BLR on DBS linked-reads yielded megabase-scale phasing with low (<0.2%) switch error rates. Of 13616 protein-coding genes phased in the GIAB benchmark set (v4.2.1), 98.6% matched the BLR phasing. In addition, large structural variants showed concordance with HPRC-HG002 reference assembly calls. Compared to diploid assembly with PacBio HiFi reads, BLR phasing was more continuous when considering switch errors. We further show that integrating long reads at low coverage (∼10×) can improve phasing contiguity and reduce switch errors in tandem repeats. When compared to Long Ranger on 10× Genomics data, BLR showed an increase in phase block N50 with low switch-error rates. For TELL-Seq and stLFR linked reads, BLR generated longer or similar phase block lengths and low switch error rates compared to results presented in the original publications. In conclusion, BLR provides a flexible workflow for comprehensive haplotype analysis of linked reads from multiple platforms., (© The Author(s) 2023. Published by Oxford University Press on behalf of Nucleic Acids Research.)

Discusses navigation in instructional hypertext programs and describes two preliminary studies that investigated concrete versus abstract representations for backward navigation. Topics include representation and hypermedia sign systems; and consistency in the forms designers use for navigation buttons. (LRW)

The future of human genomics is one that seeks to resolve the entirety of genetic variation through sequencing. The prospect of utilizing genomics for medical purposes require cost-efficient and accurate base calling, long-range haplotyping capability, and reliable calling of structural variants. Short-read sequencing has lead the development towards such a future but has struggled to meet the latter two of these needs. To address this limitation, we developed a technology that preserves the molecular origin of short sequencing reads, with an insignificant increase to sequencing costs. We demonstrate a novel library preparation method for high throughput barcoding of short reads where millions of random barcodes can be used to reconstruct megabase-scale phase blocks.

Antibody-DNA conjugates are powerful tools for DNA-assisted protein analysis. Growing usage of these methods demands efficient production of high-quality conjugates. We developed an easy and fast synthesis route yielding covalent antibody-DNA conjugates with a defined conjugation site and low batch-to-batch variability. We utilize the Z domain from protein A, containing the unnatural amino acid 4-benzoylphenylalanine (BPA) for photoaffinity labeling of the antibodies' Fc region. Z( x BPA) domains are C-terminally modified with triple-glycine (G 3 )-modified DNA-oligonucleotides via enzymatic Sortase A coupling. We show reliable modification of the most commonly used IgG's. To prove our conjugates' functionality, we detected antibody-antigen binding events in an assay called Droplet Barcode Sequencing for Protein analysis (DBS-Pro). It confirms not only retained functionality for both conjugate parts but also the potential of using DBS-Pro for quantifying protein abundances. As intermediates are easily storable and our approach is modular, it offers a convenient strategy for screening various antibody-DNA conjugates using the same starting material.

Raman spectroscopy can provide the biomolecular fingerprint of a cell in a label-free manner. Although a variety of clinical and biomedical applications have been demonstrated, the method remains largely a niche technology. The two main problems are the complexity of data acquisition and the complexity of data analysis. Generally, Raman measurements are performed manually and require a substantial amount of time. This, on the other hand, frequently results in a low number of samples and hence with questionable statistical evaluation. Here, we propose an automated high content screening Raman spectroscopy (HCS-RS) platform, which can perform a series of experiments without human interaction, significantly increasing the number of measured samples and making the measurement more reliable. The automated image processing of bright field images in combination with automatic spectral acquisition of the molecular fingerprint of cells exposed to different physiological conditions enables label-free high content screening applications. The performance of the developed HCS-RS platform is demonstrated by investigating the effect of panitumumab on SW48 and SW480 colorectal cancer cells with wild-type and mutated K-RAS, respectively, in a series of concentrations. Our result indicates that the increased content of panitumumab prohibits the activation of the MAP kinase of the colorectal cancer cells with wild-type K-RAS strongly, whereas there is no significant effect on the K-RAS mutated cells. Moreover, the relative amount of the panitumumab content present in the cells is determined from the Raman spectral information, which could be beneficial for personalized patient treatment.

Our hypothesis was that increasing the inclusion level of dried distiller's grains with solubles (DDGS) to feedlot lambs would increase growth and the inclusion of lasalocid (LAS; Bovatec, Alpharma, LLC, Bridgewater, NJ) would increase ADG and G:F, while not affecting digestibility, ruminal VFA concentration, and ruminal pH. Furthermore, we hypothesized that rations containing LAS and higher levels of DDGS would cause increased ruminal hydrogen sulfide gas (H2S) concentrations. Two hundred forty crossbred (Suffolk x Rambouillet) lambs (31.9 ± 5.87 kg BW; approximately 90 d of age) were allocated to 6 treatments in a completely randomized design with a 3 x 2 factorial arrangement of treatments. Lambs were placed into 24 feedlot pens (4 pens/treatment; 10 lambs/pen) for a 111 d finishing study. Main effects included concentration of DDGS (0, 15, or 30% DM basis) and inclusion of LAS (0 or 22.05 g/metric ton LAS) resulting in treatments of: 1) 0% DDGS without LAS (0DDGS-NL), 2) 0% DDGS with LAS (0DDGS-L), 3) 15% DDGS without LAS (15DDGS-NL), 4) 15% DDGS with LAS (15DDGS-L), 5) 30% DDGS without LAS (30DDGS-NL), and 6) 30% DDGS with LAS (30DDGS-L). Two-day weights were taken at the beginning and end of the experiment. Two-hundred-eighteen lambs (64.8 ± 7.99 kg BW) were slaughtered on d 112 at a commercial abattoir and carcass data collected. The inclusion of LAS increased (P ≤ 0.02) final BW, ADG, G:F, and HCW. As DDGS in the ration increased to 30%, DMI decreased linearly (P = 0.03) while G:F increased linearly (P = 0.03). A second study was conducted utilizing the same treatments to evaluate N and S balance, ruminal VFA and H2S concentration, and ruminal pH in 24 crossbred wethers (Suffolk x Rambouillet; 41.2 ± 12.23 kg BW). Daily urinary sulfur excretion and ruminal H2S concentration were linearly increased (P < 0.001) as DDGS increased in the ration. Total ruminal VFA concentration linearly decreased (P = 0.002) as DDGS increased in the ration. The inclusion of LAS increased (P = 0.02) ruminal pH. The results confirm our hypothesis that LAS increased overall growth and increasing DDGS increased ruminal H2S concentration but did not influence growth. We reject the hypothesis that the combined effects of LAS and DDGS would have no effect on rumen pH and VFA concentrations. [ABSTRACT FROM AUTHOR]

Stroke is the second result for death and ischemic stroke constitutes most of all stroke cases. Ischemic stroke takes place when blood clot or embolus blocks cerebral vessel and interrupts blood flow, which often leads to brain damage, permanent disability, or death. There is a 4.5-h (golden hour) treatment window to restore blood flow prior to permanent neurological impairment results. Current stroke treatments consist mechanical system or thrombolytic drug therapy to disrupt or dissolve thrombus. Promising method for stroke treatment is mechanical retrieving of thrombi employing device deployed endovascularly. Advent of smart materials has led to research fabrication of several minimally invasive endovascular devices that take advantage of new materials capabilities. One of these capabilities is shape memory, is capability of material to store temporary form, then activate to primary shape as subjected to stimuli. Shape memory polymers (SMPs) are employed as good materials for thrombectomy device fabrication. Therefore, current review presents thrombectomy device development and fabrication with SMPs. Design, performance, limitations, and in vitro or in vivo clinical results of SMP-based thrombectomy devices are identified. Review also sheds light on SMP's future outlook and recommendations for thrombectomy device application, opening a new era for advanced materials in materials science., (© 2024 The Authors. Macromolecular Rapid Communications published by Wiley‐VCH GmbH.)

Laser drop on demand jetting of Cu-base braze droplets was proven a suitable method for joining wires to electrode structures of electronic devices, particularly if the electrical contacts need to withstand high thermal loads. During joining, a braze preform of 600 µm diameter is placed inside a capillary, molten by a laser pulse and subsequently ejected from the capillary by inert gas overpressure similarly to conventional solder ball bumping processes. However, since the liquidus temperature of the used braze material of 990 °C is about 760 °C higher than of standard Sn-based solders used in electronics packaging, the system technology was modified significantly to enable jetting of CuSn alloys. In particular, the beam source emits a five times higher optical output power than standard machines designed for processing Sn-based solders. In addition, a modified capillary made from technical ceramic was machined, to withstand the significantly higher heating- and cooling rates during the process. In order to understand the influence of capillary geometry on droplet detachment, and flight trajectory, two capillary geometries were machined applying a picosecond laser ablation process. Subsequently, stereoscopic high speed videos of droplet detachment and flight phase were analyzed. Using this approach it is possible, to determine droplet flight trajectories, velocities and lateral positional deviations in dependency of relative inert gas overpressure inside the machining head, pulse power and capillary geometry. The findings indicate a significant influence of the capillary geometry and the applied overpressure on the droplet flight trajectory, whereas the role of the laser pulse power seems neglectable.

Background: The small molecule pifithrin-μ reversibility inhibits the mitochondrial pathway of apoptosis. The neuronal effects of pifithrin-μ applied after cardiac arrest are unknown. We hypothesized that pifithrin-μ reduces neuronal damage in the most vulnerable brain region, the hippocampus, after cardiac arrest., Methods: In two randomized controlled series we administered pifithrin-μ or control in 109 rats resuscitated after 8 or 10 min of cardiac arrest. Neuronal damage was blindly assessed with histology (Fluoro Jade B: FJB, cresyl violet: CV) in the most vulnerable brain region (CA1 segment of hippocampus) and with a series of neurobehavioral tests (Open Field Task, Tape-Removal Test, Morris Water Maze test). Mixed ANOVA was used to combine both series, simple comparisons were done with t tests or Mann-Whitney U test., Results: Pifithrin-μ reduced the number of degenerating, FJB-positive neurons by 25% (mixed ANOVA p group = 0.014). This was more prominent after 8 min cardiac arrest (8 min arrest pifithrin-μ 94 ± 47 vs control 128 ± 37; n = 11 each; 10 min arrest pifithrin-μ 78 ± 44, n = 15 vs control 101 ± 31, n = 18; p group* arrest length interaction = 0.622). The reduction of ischemic CV-positive neurons in pifithrin-μ animals was not significant (ANOVA p group = 0.063). No significant group differences were found in neurobehavioral testing., Conclusion: Temporarily inhibition of apoptosis with pifithrin-μ after cardiac arrest decreases the number of injured neurons in the CA1 segment of hippocampus in a cardiac arrest rat model, without clinical correlate. Further studies should elucidate the role of this neuroprotective agent in different settings and with longer cardiac arrest.