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- Abbasi Aval, Negar, et al.
(författare)
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Assessing the Layer-by-Layer Assembly of Cellulose Nanofibrils and Polyelectrolytes in Pancreatic Tumor Spheroid Formation
- 2023
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Ingår i: Biomedicines. - : MDPI AG. - 2227-9059. ; 11:11
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Tidskriftsartikel (refereegranskat)abstract
- Three-dimensional (3D) tumor spheroids are regarded as promising models for utilization as preclinical assessments of chemo-sensitivity. However, the creation of these tumor spheroids presents challenges, given that not all tumor cell lines are able to form consistent and regular spheroids. In this context, we have developed a novel layer-by-layer coating of cellulose nanofibril–polyelectrolyte bilayers for the generation of spheroids. This technique builds bilayers of cellulose nanofibrils and polyelectrolytes and is used here to coat two distinct 96-well plate types: nontreated/non-sterilized and Nunclon Delta. In this work, we optimized the protocol aimed at generating and characterizing spheroids on difficult-to-grow pancreatic tumor cell lines. Here, diverse parameters were explored, encompassing the bilayer count (five and ten) and multiple cell-seeding concentrations (10, 100, 200, 500, and 1000 cells per well), using four pancreatic tumor cell lines—KPCT, PANC-1, MiaPaCa-2, and CFPAC-I. The evaluation includes the quantification (number of spheroids, size, and morphology) and proliferation of the produced spheroids, as well as an assessment of their viability. Notably, our findings reveal a significant influence from both the number of bilayers and the plate type used on the successful formation of spheroids. The novel and simple layer-by-layer-based coating method has the potential to offer the large-scale production of spheroids across a spectrum of tumor cell lines.
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| 5. |
- Abbasi Aval, Negar, 1986-
(författare)
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Utilizing Biopolymers in 3D Tumor Modeling and Tumor Diagnosis
- 2023
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Cancer represents a significant global public health challenge and ranks as the second mostcommon cause of death in the United States. The onset of cancer entails an initial phasewhere cells lose their polarity and disconnect from the normal basement membrane, allowingthem to form distinct three-dimensional (3D) configurations that interact with adjacent cellsand the surrounding microenvironment. Cells grown in 2D monolayers demonstrate differentgene expression patterns and different activation of signaling pathways compared to cellscultivated within the natural structure of tumor tissue of the same cell type. Multicellulartumor spheroids (MCTS) are extensively investigated as a well-studied model of organotypiccancer. These spheroids are formed by tumor cells, either alone or in combination with othercell types, and they can be created with or without the application of supportive scaffolds.The MCTSs are also considered promising models for preclinical assessments of chemosensitivity.However, the creation of these tumor spheroids presents challenges, as not alltumor cell lines can consistently form regular spheroids.Cellulose nanofibrils (CNF) have become essential as a sustainable and environmentallyfriendly material. For example, thin films, with inherent mechanical properties, and flexibility,offer versatility across various applications. Also known for its biocompatibility and non-toxicnature, native CNF is a natural option to use. Its fibrous structure closely mimics the collagenmatrix in human tissue, showing potential as an effective scaffold for 3D cell culture. In thisregard, an innovative Layer-by-Layer (LbL) coating technique using CNF-polyelectrolytebilayers was investigated to generate spheroids. This method constructs bilayers of CNFand polyelectrolytes and can coat various surfaces. In this thesis, the first focus was ondemonstrating the spheroid formation capability using low molecular weight polyelectrolytesin LbL assembly. Secondly, an investigation was conducted involving embedding of LbLgrownspheroids in a decellularized extracellular matrix (ECM) aiming to determine howECM, possessing suitable mechanical characteristics, could influence the cancer stem celltraits in spheroids. Thirdly, the thesis demonstrated the utilization of LbL for capturing andreleasing of circulating tumor cells. Lastly, the shift from using low molecular weightpolyelectrolytes in the LbL assembly to high molecular weight counterparts and analyzingthe differences in spheroid formation abilities to assess the underlying differences inmolecular weights of the polyelectrolytes was explored. All-in-all, employing the CNF-basedLbL surface coating strategy explored in the thesis has proven to be promising for thedevelopment of spheroid models closely resembling in vivo conditions and holds significantpotential for applications in drug development.
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| 7. |
- Akhtar, Ahmad Saleem, et al.
(författare)
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A portable and low-cost centrifugal microfluidic platform for multiplexed colorimetric detection of protein biomarkers
- 2023
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Ingår i: Analytica Chimica Acta. - : Elsevier BV. - 0003-2670 .- 1873-4324. ; 1245
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Tidskriftsartikel (refereegranskat)abstract
- Cytokines play a very important role in our immune system by acting as mediators to put up a coordinated defense against foreign elements in our body. Elevated levels of cytokines in the body can signal to an ongoing response of the immune system to some abnormality. Thus, the quantification of a panel of cytokines can provide valuable information regarding the diagnosis of specific diseases and state of overall health of an individual. Conventional Enzyme Linked Immunosorbent Assay (ELISA) is the gold-standard for quantification of cytokines, however the need for trained personnel and expensive equipment limits its application to centralized laboratories only. In this context, there is a lack of simple, low-cost and portable devices which can allow for quantification of panels of cytokines at point-of-care and/or resource limited settings.Here, we report the development of a versatile, low-cost and portable bead-based centrifugal microfluidic platform allowing for multiplexed detection of cytokines with minimal hands-on time and an integrated colorimetric signal readout without the need for any external equipment. As a model, multiplexed colorimetric quantification of three target cytokines i.e., Tumor necrosis factor alpha (TNF-α), Interferon gamma (IFN-γ) and Interleukin-2 (IL-2) was achieved in less than 30 min with limits of detection in ng/mL range. The developed platform was further evaluated using spiked-in plasma samples to test for matrix interference. The ease of use, low-cost and portability of the developed platform highlight its potential to serve as a sample-to-answer solution for detection of cytokine panels in resource limited settings.
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| 8. |
- Akhtar, Ahmad Saleem, et al.
(författare)
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An integrated centrifugal microfluidic platform for multiplexed colorimetric immunodetection of protein biomarkers in resource-limited settings
- 2021
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Ingår i: Proceedings MicroTAS 2021 - 25th International Conference on Miniaturized Systems for Chemistry and Life Sciences. - : Chemical and Biological Microsystems Society. ; , s. 947-948
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Konferensbidrag (refereegranskat)abstract
- The up- and down- regulation of inflammatory biomarkers such as cytokines can be indicative of several diseases such as primary cancers and/or metastatic tumors, as well as less serious conditions. For point-of-care clinical applications, the detection of these biomarkers requires a combination of a sensitive assay and multiplexing capabilities, together with fit-for-purpose signal transduction strategies. Here, we report the development of a versatile and cost-effective integrated centrifugal microfluidic platform compatible with resource-limited settings using nanoporous microbeads for immunoaffinity-based profiling of cytokines. With an automated colorimetric readout at the end, the platform allows for profiling of cytokines in < 30 mins.
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- Akhtar, Ahmad Saleem, et al.
(författare)
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Centrifugal microfluidic platform comprising an array of bead microcolumns for the multiplexed colorimetric quantification of inflammatory biomarkers at the point-of-care
- 2019
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Ingår i: 23rd International Conference on Miniaturized Systems for Chemistry and Life Sciences, MicroTAS 2019. - : Chemical and Biological Microsystems Society. ; , s. 1230-1231
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Konferensbidrag (refereegranskat)abstract
- The detection of panels of inflammatory biomarkers such as cytokines has potential for the rapid and specific diagnostic of several devastating diseases such as primary cancers and/or metastatic tumors, as opposed to less serious conditions. For point-of-care clinical applications, the detection of these biomarkers requires a combination of pg/mL sensitivities and multiplexing capabilities, coupled with fit-for-purpose signal transduction strategies. Here, we report the development of a versatile centrifugal microfluidic platform combined with nanoporous microbeads for immunoaffinity-based profiling of cytokines. The device allows sample and analyte multiplexing and detection limits below 1 ng/mL were achieved within 30 minutes, using colorimetric detection.
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| 10. |
- Akhtar, Ahmad Saleem
(författare)
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Centrifugal microfluidics-based point of care diagnostics at resource limited settings
- 2023
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Advancements in medical diagnostics have allowed us to understand the underlying mechanism and treat the root cause for many diseases which had been causing morbidity and mortality up until this point in human history. Furthermore, many of the standard diagnostic procedures have now been transformed to provide answers at or near the point-of-care. However, the effects of these positive developments have not trickled down to the parts of our society which are considered underdeveloped and lack the necessary infrastructure and facilities. Diagnostics in such resource limited settings still lag behind and fail to provide the requisite healthcare. In order to translate the diagnostic solutions designed for central laboratories to resource limited settings, there are certain challenges that need to be addressed, such as portability, reduction in cost and ease-of-use, while keeping the sensitivity and specificity at the required level. The work presented in this thesis focuses on addressing some of these issues by using microfluidics to develop diagnostic platforms that are suitable to be used in resource limited settings. In paper I, a very low-cost and simple centrifugal microfluidic platform was developed to be used in settings which do not have a reliable supply of electricity. The platform uses a smartphone as a source of power and the sensors of the phone for speed control.In paper II, a portable and low-cost diagnostic platform was developed for multiplexed detection of biomarkers based on centrifugal microfluidics. The platform uses colorimetric detection and a simple readout method which does not require a spectrophotometer for quantification.In paper III, a platform was developed for COVID-19 diagnostics which combines centrifugal microfluidics with a novel bead-based strategy for signal enhancement. The platform uses fluorescent detection with a smartphone readout and has the capability to process up to 20 samples at the same time.In paper IV, as a follow up of paper III, a more advanced platform was developed for COVID-19 diagnostics which allows the operator to carry out nucleic acid amplification in a completely automated manner, from adding the sample to getting the final result.In paper V, an alternative method for detection of SARS-CoV-2 was developed using electrochemical biosensing. This work combines the electrochemical technique with a flexible printed circuit board for a rapid, amplification-free and label-free detection of target SARS-CoV-2 sequences.
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