| 1. |
- Baldwin, Lydia, et al.
(författare)
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Development of a dual-flow tissue perfusion device for modeling the gastrointestinal tract-brain axis
- 2023
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Ingår i: Biomicrofluidics. - 1932-1058. ; 17:5
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Tidskriftsartikel (refereegranskat)abstract
- Despite the large number of microfluidic devices that have been described over the past decade for the study of tissues and organs, few have become widely adopted. There are many reasons for this lack of adoption, primarily that devices are constructed for a single purpose or because they are highly complex and require relatively expensive investment in facilities and training. Here, we describe a microphysiological system (MPS) that is simple to use and provides fluid channels above and below cells, or tissue biopsies, maintained on a disposable, poly(methyl methacrylate), carrier held between polycarbonate outer plates. All other fittings are standard Luer sizes for ease of adoption. The carrier can be coated with cells on both sides to generate membrane barriers, and the devices can be established in series to allow medium to flow from one cell layer to another. Furthermore, the carrier containing cells can be easily removed after treatment on the device and the cells can be visualized or recovered for additional off-chip analysis. A 0.4 mu m membrane with cell monolayers proved most effective in maintaining separate fluid flows, allowing apical and basal surfaces to be perfused independently. A panel of different cell lines (Caco-2, HT29-MTX-E12, SH-SY5Y, and HUVEC) were successfully maintained in the MPS for up to 7 days, either alone or on devices connected in series. The presence of tight junctions and mucin was expressed as expected by Caco-2 and HT-29-MTX-E12, with Concanavalin A showing uniform staining. Addition of Annexin V and PI showed viability of these cells to be >80% at 7 days. Bacterial extracellular vesicles (BEVs) produced by Bacteroides thetaiotaomicron and labeled with 1,1 '-dioctadecyl-3,3,3 ',3 '-tetramethylindocarbo-cyanine perchlorate (DiD) were used as a model component of the human colonic microbiota and were visualized translocating from an apical surface containing Caco-2 cells to differentiated SH-SY5Y neuronal cells cultured on the basal surface of connected devices. The newly described MPS can be easily adapted, by changing the carrier to maintain spheroids, pieces, or slices of biopsy tissue and joined in series to study a variety of cell and tissue processes. The cell layers can be made more complex through the addition of multiple cell types and/or different patterning of extracellular matrix and the ability to culture cells adjacent to one another to allow study of cell:cell transfer, e.g., passive or active drug transfer, virus or bacterial entry or BEV uptake and transfer.
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| 2. |
- Barry, Antonia, et al.
(författare)
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Investigating the effects of arginine methylation inhibitors on microdissected brain tumour biopsies maintained in a miniaturised perfusion system
- 2023
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Ingår i: Lab on a Chip. - 1473-0197 .- 1473-0189. ; 23:11, s. 2664-2682
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Tidskriftsartikel (refereegranskat)abstract
- Arginine methylation is a post-translational modification that consists of the transfer of one or two methyl (CH3) groups to arginine residues in proteins. Several types of arginine methylation occur, namely monomethylation, symmetric dimethylation and asymmetric dimethylation, which are catalysed by different protein arginine methyltransferases (PRMTs). Inhibitors of PRMTs have recently entered clinical trials to target several types of cancer, including gliomas (NCT04089449). People with glioblastoma (GBM), the most aggressive form of brain tumour, are among those with the poorest quality of life and likelihood of survival of anyone diagnosed with cancer. There is currently a lack of (pre)clinical research on the possible application of PRMT inhibitors to target brain tumours. Here, we set out to investigate the effects of clinically-relevant PRMT inhibitors on GBM biopsies. We present a new, low-cost, easy to fabricate perfusion device that can maintain GBM tissue in a viable condition for at least eight days post-surgical resection. The miniaturised perfusion device enables the treatment of GBM tissue with PRMT inhibitors ex vivo, and we observed a two-fold increase in apoptosis in treated samples compared to parallel control experiments. Mechanistically, we show thousands of differentially expressed genes after treatment, and changes in the type of arginine methylation of the RNA binding protein FUS that are consistent with hundreds of differential gene splicing events. This is the first time that cross-talk between different types of arginine methylation has been observed in clinical samples after treatment with PRMT inhibitors.
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| 3. |
- Changtor, Phanupong, 1994-, et al.
(författare)
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Integration of IFAST-based nucleic acid extraction and LAMP for on-chip rapid detection of Agroathelia rolfsii in soil
- 2024
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Ingår i: Biosensors & bioelectronics. - 0956-5663 .- 1873-4235. ; 250
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Tidskriftsartikel (refereegranskat)abstract
- Agroathelia rolfsii (A. rolfsii) is a fungal infection and poses a significant threat to over 500 plant species worldwide. It can reduce crop yields drastically resulting in substantial economic losses. While conventional detection methods like PCR offer high sensitivity and specificity, they require specialized and expensive equipment, limiting their applicability in resource -limited settings and in the field. Herein, we present an integrated workflow with nucleic acid extraction and isothermal amplification in a lab -on -a -chip cartridge based on immiscible filtration assisted by surface tension (IFAST) to detect A. rolfsii fungi in soil for point -of -need application. Our approach enabled both DNA extraction of A. rolfsii from soil and subsequent colorimetric loop -mediated isothermal amplification (LAMP) to be completed on a single chip, termed IFAST-LAMP. LAMP primers targeting ITS region of A. rolfsii were newly designed and tested. Two DNA extraction methods based on silica paramagnetic particles (PMPs) and three LAMP assays were compared. The best -performing assay was selected for on -chip extraction and detection of A. rolfsii from soil samples inoculated with concentrations of 3.75, 0.375 and 0.0375 mg fresh weight per 100-g soil (%FW). The full on -chip workflow was achieved within a 1-h turnaround time. The platform was capable of detecting as low as 3.75 %FW at 2 days after inoculation and down to 0.0375 %FW at 3 days after inoculation. The IFAST-LAMP could be suitable for field -applicability for A. rolfsii detection in low -resource settings.
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| 4. |
- Ngamsom, Bongkot, et al.
(författare)
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A sample-to-answer COVID-19 diagnostic device based on immiscible filtration and CRISPR-Cas12a-assisted detection
- 2022
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Ingår i: Talanta Open. - : Elsevier BV. - 2666-8319. ; 6
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Tidskriftsartikel (refereegranskat)abstract
- In response to the ongoing coronavirus disease 2019 (COVID-19) pandemic and disparities of vaccination coverage in low-and middle-income countries, it is vital to adopt a widespread testing and screening programme, combined with contact tracing, to monitor and effectively control the infection dispersion in areas where medical resources are limited. This work presents a lab-on-a-chip device, namely ‘IFAST-LAMP-CRISPR’, as an affordable, rapid and high-precision molecular diagnostic means for detection of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2). The herein proposed ‘sample-to-answer’ platform integrates RNA extraction, amplification and molecular detection with lateral flow readout in one device. The microscale dimensions of the device containing immiscible liquids, coupled with the use of silica paramagnetic beads and guanidine hydrochloride, streamline sample preparation (including RNA extraction, concentration and purification) in 15 min with minimal hands-on steps. The pre-amplification in combination with CRISPR-Cas12a detection assays targeting the nucleoprotein (N) gene achieved visual identification of ≥ 470 copies mL−1 genomic SARS-CoV-2 samples in 45 min. On-chip assays showed the ability to isolate and detect SARS-CoV-2 RNA from 100 genome copies mL−1 of replication-deficient viral particles in 1 h. This simple, affordable and integrated platform demonstrated a visual, faster, and yet specificity- and sensitivity-comparable alternative to the costly gold-standard reverse transcription-polymerase chain reaction (RT-PCR) assay, requiring only a simple heating source. Initial testing illustrates the platform viability both on nasopharyngeal swab and saliva samples collected using the easily accessible Swan-brand cigarette filter, providing a complete workflow for COVID-19 diagnostics in low-resource settings.
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| 5. |
- Oday, Jwan, et al.
(författare)
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Development and validation of spectrophotometric method and paper-based microfluidic devices for the quantitative determination of Amoxicillin in pure form and pharmaceutical formulations
- 2024
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Ingår i: Heliyon. - 2405-8440. ; 10:3
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Tidskriftsartikel (refereegranskat)abstract
- There is a growing need for easy-to-use, low cost and portable quantitative assays to determine active pharmaceutical ingredients in the pharmaceutical industry. Here, we developed a batch spectrophotometric method and a method employing a paper-based microfluidic device for the estimation of Amoxicillin (AMX) in pure solution and pharmaceutical preparations. The detection depends on the coupling reaction of Amoxicillin with diazotized sulfadimidine (DSDM) in an alkaline medium. The yellow azo dye reaction product was measured at λmax 425 nm and linearity was observed from 2 to 30 mg L−1 with a detection limit of 0.32 mg L−1 and a quantification limit of 1.2 mg L−1 was found. The reaction was then transferred onto the paper-based microfluidic device and a plateau change in color intensity was found above 10 mg L−1. Thus, the paper-based microfluidic device can be applied for the semi-quantitative determination of Amoxicillin in pure solution and commercial pharmaceutical products for rapid screening.
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| 6. |
- Rodriguez Mateos, Pablo, 1995-, et al.
(författare)
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Integrated microscale immiscible phase extraction and isothermal amplification for colorimetric detection of Neisseria gonorrhoeae
- 2023
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Ingår i: Analytical and Bioanalytical Chemistry. - 1618-2642 .- 1618-2650. ; 415:21, s. 5129-5137
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Tidskriftsartikel (refereegranskat)abstract
- Gonorrhea is the second most common sexually transmitted infection (STI) with around 87 million cases worldwide estimated in 2016 by the World Health Organization. With over half of the cases being asymptomatic, potential life-threatening complications and increasing numbers of drug-resistant strains, routine monitoring of prevalence and incidence of infections are key preventive measures. Whilst gold standard qPCR tests have excellent accuracy, they are neither affordable nor accessible in low-resource settings. In this study, we developed a lab-on-a-chip platform based on microscale immiscible filtration to extract, concentrate and purify Neisseria gonorrhoeae DNA with an integrated detection assay based on colorimetric isothermal amplification. The platform was capable of detecting as low as 500 copies/mL from spiked synthetic urine and showed no cross-reactivity when challenged with DNAs from other common STIs. The credit card-size device allows DNA extraction and purification without power or centrifuges, and the detection reaction only needs a low-tech block heater, providing a straightforward and visual positive/negative result within 1 h. These advantages offer great potential for accurate, affordable and accessible monitoring of gonorrhea infection in resource-poor settings.
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| 7. |
- Rodriguez Mateos, Pablo, 1995-, et al.
(författare)
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Microscale immiscible phase magnetic processing for bioanalytical applications
- 2023
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Ingår i: TrAC. Trends in analytical chemistry. - : Elsevier BV. - 0165-9936 .- 1879-3142. ; 158
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Forskningsöversikt (refereegranskat)abstract
- Bioanalytical processes such as ligand-binding assays, nucleic acid amplification tests and cancer cell identification methods require multiple steps including sampling, transport, sample preparation, chemical reaction/s, detection and interpretation. These necessitate trained personnel and laboratory infrastructure, and are often time-consuming and labor-intensive. Integrated platforms capable of combining the aforementioned steps are generally challenging to realize. Approaches employing immiscible phases constrained in the microscale and paramagnetic particles have emerged as pathways to extract proteins, nucleic acids and whole cells from complex matrices via affinity purification and magnetic actuation. In recent years, further steps for detection and readout of these bioanalytes have been incorporated, providing versatile lab-on-a-chip platforms for multiple integrated bioanalytical workflows and with great potential as point-of-need devices. In this manuscript, we review and summarize the different strategies employed and lay out further avenues for development and implementation.
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