ARTICLE | doi:10.20944/preprints202311.1302.v1
Subject: Biology And Life Sciences, Life Sciences Keywords: glycosylation; monocloncal antobody; immunoglobulin
Online: 21 November 2023 (10:02:46 CET)
Glycosylation is a common post-translation modification present on majority of eukaryotic proteins. Therefore, it is dependent on the host cell line and affected by the cell culture and purification process. Additionally, glycosylation of therapeutic biologics is known to have a profound impact on binding and downstream effector functions. Hence, glycosylation is increasingly considered a critical quality attribute (CQA) of biopharmaceutical drugs and has been showed to affect the safety and efficacy of these molecules. Therefore, regulatory agencies have adopted a Quality by Design (QbD) approach for glycan monitoring at various steps with stringent controls to ensure efficacy and safety. This requires multiple, orthogonal fit-for-purpose tools to study the process. One key unmet need in downstream organizations of large pharmaceutical companies is a real-time, rapid glycan detection and quantification tool to guide purification/expression process. To meet this need, we have developed a selective, colorimetric assay which selectively stains sugar moieties (glycosylated IgGs) which appears as magenta bands on an SDS-PAGE resolved gel. The selectivity arises because the method leverages stereochemical differences of hydroxyl groups between amino acids and carbohydrates to selectively stain the primary alcohol in carbohydrates but not in amino acids. Specifically, our method selectively oxidizes the ‘cis-diol’ group in carbohydrates to aldehydes and stains the aldehydes on an SDS-PAGE gel by Schiff’s reaction. Our method is semi-quantitative and has comparable sensitivity to Coomassie with LoD of around 50ng. The method can detect glycosylated IgGs produced in multiple cell lines (including HEK and CHO), thereby enabling us to study batch-to-batch variability and can critically stain both N-linked and O-linked glycosylation. We also show that the assay can be used as a downstream purification guide for process analytics following ion-exchange and hydrophobic interaction chromatography.
ARTICLE | doi:10.20944/preprints201711.0110.v1
Subject: Biology And Life Sciences, Biology And Biotechnology Keywords: Fluorescent reporter; E2-Crimson; mouse embryonic stem cells; knock-in; in vivo imaging
Online: 16 November 2017 (17:46:53 CET)
Far-red fluorescent reporter genes can be used for tracking cells non-invasively in vivo using fluorescence imaging. Here, we investigate the effectiveness of the far-red fluorescent protein, E2-Crimson (E2C), for tracking mouse embryonic cells (mESCs) in vivo following subcutaneous administration into mice. Using a knock-in strategy, we introduced E2C into the Rosa26 locus of an E14-Bra-GFP mESC line, and after confirming that the E2C had no obvious effect on the phenotype of the mESCs, we injected them into mice and imaged them over 9 days. The results showed that fluorescence intensity was weak, and cells could only be detected when injected at high densities. Furthermore, intensity peaked on day 4 and then started to decrease, despite the fact that tumour volume continued to increase beyond day 4. Histopathological analysis showed that although E2C fluorescence could barely be detected in vivo at day 9, analysis of frozen sections indicated that all mESCs within the tumours continued to express E2C. We hypothesise that the decrease in fluorescence intensity in vivo was probably due to the fact that the mESC tumours became more vascular with time, thus leading to increased absorbance of E2C fluorescence by haemoglobin. We conclude that the E2C reporter has limited use for tracking cells in vivo, at least when introduced as a single copy into the Rosa26 locus.
ARTICLE | doi:10.20944/preprints202307.1991.v1
Subject: Computer Science And Mathematics, Artificial Intelligence And Machine Learning Keywords: textile fabrics; color fastness; digital grading; spectral reconstruction; BP neural network
Online: 28 July 2023 (10:42:44 CEST)
To digital grade the color fastness of fabrics after rubbing, an automatic grading method based on spectral reconstruction technology and BP neural network was proposed. Firstly, the modeling samples are prepared by rubbing the fabrics according to the ISO standard of 105-X12. Then, to comply with visual rating standards for color fastness, the modeling samples are professionally graded to obtain the visual rating result. After that, a digital camera is used to capture digital images of the modeling samples inside a closed and uniform lighting box, and the color data values of the modeling samples are obtained through spectral reconstruction technology. Finally, the color fastness prediction model for rubbing was constructed using the modeling samples data and BP neural network. The color fastness level of the testing samples was predicted using the prediction model, and the prediction results were compared with the existing color difference conversion method and curve fitting method. Experimental show that the prediction model of fabric color fastness can be better constructed using the BP neural network, where the root-mean-square error of the prediction for the training sample is 0.30, and the root-mean-square error of the prediction for the test sample is 0.25. The overall performance of the method is slightly better than the color difference conversion method, and it is significantly outperforming the curve fitting method. It can be seen that the digital rating method of fabric color fastness to rubbing based on spectral reconstruction and BP neural network has a high consistency with the visual evaluation, which will help for the automatic color fastness grading.
ARTICLE | doi:10.20944/preprints201705.0008.v1
Subject: Chemistry And Materials Science, Surfaces, Coatings And Films Keywords: polyester fabrics; surface modification; sulfuric acid; polypyrrole; deposition; conductivity
Online: 1 May 2017 (08:48:17 CEST)
A simple and effective surface modification of polyester fabrics with sulfuric acid to improve the interfacial deposition of polypyrrole was presented in our work. A range of sulfuric acid concentrations were analyzed by studying water contact angle. Effect of sulfuric acid modification on the deposition of polypyrrole was investigated by sheet resistance and color depth of fabric samples. Polyester fabrics coated with polypyrrole layer were confirmed by scanning electron microscopy (SEM), X-ray photoelectron spectroscopy (XPS), X-Ray diffraction spectra (XRD), Fourier transform infrared spectroscopy (FTIR). XPS showed that sulphur containing functional groups were obviously appeared on the polyester fiber surface after modification, which were advantageous to promote the deposition of polypyrrole onto polyester fabrics. The improved deposition increased electrical conductivity of fabric samples.
REVIEW | doi:10.20944/preprints201812.0156.v1
Subject: Biology And Life Sciences, Ecology, Evolution, Behavior And Systematics Keywords: Marine microorganisms, marine fungi, algae, marine plants, marine invertebrates, marine vertebrates, genome, metagenome
Online: 12 December 2018 (15:44:09 CET)
With the continuing development of sequencing technology, genomics has been applied in a variety of biological research areas. In particular, the application of genomics to marine species, which boast a high diversity, promises great scientific and industrial potential. Significant progress has been made in marine genomics especially over the past few years. Consequently, BGI, leveraging its prominent contributions in genomics research, established BGI-Qingdao, an institute specifically aimed at exploring marine genomics. In order to accelerate marine genomics research and related applications, BGI-Qingdao initiated the International Conference on Genomics of the Ocean (ICG-Ocean) to develop international collaborations and establish a focused and coherent global research plan. Last year, the first ICG-Ocean conference was held in Qingdao, China, during which 47 scientists in marine genomics from all over the world reported on their research progress to an audience of about 300 attendees. This year, we would like to build on that success, drafting a report on marine genomics to draw global attention to marine genomics. We summarized the recent progress, proposed future directions, and we would like to enable additional profound insights on marine genomics. Similar to the annual report on plant and fungal research by Kew Gardens, and the White Paper of ethical issues on experimental animals, we hope our first report on marine genomics can provide some useful insights for researchers, funding agencies as well as industry, and that future versions will expand upon the foundation established here in both breadth and depth of knowledge.This report summarizes the recent progress in marine genomics in six parts including: marine microorganisms, marine fungi, marine algae and plants, marine invertebrates, marine vertebrates and genomics-based applications.