| 1. |
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- 2019
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Tidskriftsartikel (refereegranskat)
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| 2. |
- Björkesten, Johan, et al.
(författare)
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Rolling circle amplification reporters – a general tool to simplify molecular detections
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Annan publikation (övrigt vetenskapligt/konstnärligt)abstract
- Methods to detect biomolecules via rolling circle amplification (RCA), for example padlock probes and proximity ligation assay (PLA) tend to be complex and include several reaction steps. Herein, we evaluated a new tool for the toolbox of RCA-based detection methods - RCA Reporters. Briefly, RCA Reporters represent inert DNA structures that, upon contact with a specific nucleic acid sequence, unravel via a highly specific strand displacement process to initiate local enzyme-assisted RCA reactions. The RCA Reporters can be used to directly detect ssDNA or RNA in a sample, or proteins via oligonucleotide-conjugated antibodies. The reagents can also enable faster RCA reactions or extremely selective genotyping of RCA products with repeated copies of a target sequence through a majority-vote mechanism. Further amplification of ongoing RCA reactions via RCA Reporters can allow efficient digital enumeration of single molecules via flow cytometry, with potential for simple and highly accurate molecular counting assays. The intrinsic simplicity of RCA Reporter also renders them attractive for applications at the point of care.
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| 3. |
- Blokzijl, Andries, et al.
(författare)
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Elevated Levels of SOX10 in Serum from Vitiligo and Melanoma Patients, Analyzed by Proximity Ligation Assay
- 2016
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Ingår i: PLOS ONE. - : Public Library of Science (PLoS). - 1932-6203. ; 11:4
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Tidskriftsartikel (refereegranskat)abstract
- BackgroundThe diagnosis of malignant melanoma currently relies on clinical inspection of the skin surface and on the histopathological status of the excised tumor. The serum marker S100B is used for prognostic estimates at later stages of the disease, but analyses are marred by false positives and inadequate sensitivity in predicting relapsing disorder.ObjectivesTo investigate SOX10 as a potential biomarker for melanoma and vitiligo.MethodsIn this study we have applied proximity ligation assay (PLA) to detect the transcription factor SOX10 as a possible serum marker for melanoma. We studied a cohort of 110 melanoma patients. We further investigated a second cohort of 85 patients with vitiligo, which is a disease that also affects melanocytes.ResultsThe specificity of the SOX10 assay in serum was high, with only 1% of healthy blood donors being positive. In contrast, elevated serum SOX10 was found with high frequency among vitiligo and melanoma patients. In patients with metastases, lack of SOX10 detection was associated with treatment benefit. In two responding patients, a change from SOX10 positivity to undetectable levels was seen before the response was evident clinically.ConclusionsWe show for the first time that SOX10 represents a promising new serum melanoma marker for detection of early stage disease, complementing the established S100B marker. Our findings imply that SOX10 can be used to monitor responses to treatment and to assess if the treatment is of benefit at stages earlier than what is possible radiologically.
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| 4. |
- Chen, Dong Lei, 1985-, et al.
(författare)
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A partial lower jaw of a tetrapod from "Romer's Gap"
- 2018
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Ingår i: Earth and environmental science transactions of the Royal Society of Edinburgh. - 1755-6910 .- 1755-6929. ; 108:1, s. 55-65
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Tidskriftsartikel (refereegranskat)abstract
- The first half of the Mississippian or Early Carboniferous (Tournaisian to mid-Visean), an interval of about 20 million years, has become known as "Romer's Gap" because of its poor tetrapod record. Recent discoveries emphasise the differences between pre-"Gap" Devonian tetrapods, unambiguous stem-group members retaining numerous "fish" characters indicative of an at least partially aquatic lifestyle, and post-"Gap" Carboniferous tetrapods, which are far more diverse and include fully terrestrial representatives of the main crown-group lineages. It seems that "Romer's Gap" coincided with the cladogenetic events leading to the origin of the tetrapod crown group. Here, we describe a partial right lower jaw ramus of a tetrapod from the late Tournaisian or early Visean of Scotland. The large and robust jaw displays a distinctive character combination, including a significant mesial lamina of the strongly sculptured angular, an open sulcus for the mandibular lateral line, a non-ossified narrow Meckelian exposure, a well-defined dorsal longitudinal denticle ridge on the prearticular, and a mesially open adductor fossa. A phylogenctic analysis places this specimen in a trichotomy with Crassigyrinus and baphetids + higher tetrapods in the upper part of the tetrapod stem group, above Whatcheeria, Pederpes, Ossinodus, Sigournea and Greererpeton. It represents a small but significant step in the gradual closure of "Romer's Gap".
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| 5. |
- Chen, Dong Lei, 1985-, et al.
(författare)
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Development of cyclic shedding teeth from semi-shedding teeth : the inner dental arcade of the stem osteichthyan Lophosteus
- 2017
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Ingår i: Royal Society Open Science. - : ROYAL SOC. - 2054-5703. ; 4:5
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Tidskriftsartikel (refereegranskat)abstract
- The numerous cushion-shaped tooth-bearing plates attributed to the stem-group osteichthyan Lophosteus superbus, which are argued here to represent the ancient form of inner dental arcade, display a unique and presumably primitive way of tooth shedding by basal hard tissue resorption. They carry regularly spaced, recumbent, gently recurved teeth arranged in transverse tooth files that diverge towards the lingual margin of the cushion. Three-dimensional (3D) reconstruction from propagation phase contrast synchrotron microtomography (PPC-SRμCT) reveals remnants of the first-generation teeth embedded in the basal plate that have never been discerned in any taxa. These teeth were shed by semi-basal resorption with the periphery of their bases retained as dentine rings. The rings are highly overlapped, which evidences tooth shedding prior to adding the next first-generation tooth. Later teeth at the same sites underwent cyclical replacing and shedding through basal resorption, producing stacks of buried resorption surfaces separated by bone of attachment. The number and spatial arrangement of resorption surfaces elucidates that basal resorption of replacement teeth had taken place at the older tooth sites before the addition of the youngest first-generation teeth at the lingual margin. Thus the replacement tooth buds cannot have been generated by a single permanent dental lamina, but must have arisen either from successional dental laminae associated with the predecessor teeth, or directly from the dental epithelium of these teeth. The virtual histological dissection of these Late Silurian microfossils broadens our understanding of the development of the gnathostome dental systems and the acquisition of the osteichthyan-type of tooth replacement.
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| 6. |
- Chen, Lei, 1985-
(författare)
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Molecular Tools for Biomarker Detection
- 2017
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- The advance of biological research promotes the emerging of new methods and solutions to answer the biological questions. This thesis describes several new molecular tools and their applications for the detection of genomic and proteomic information with extremely high sensitivity and specificity or simplify such detection procedures without compromising the performance.In paper I, we described a general method namely super RCA, for highly specific counting of single DNA molecules. Individual products of a range of molecular detection reactions are magnified to Giga-Dalton levels that are easily detected for counting one by one, using methods such as low-magnification microscopy, flow cytometry, or using a mobile phone camera. The sRCA-flow cytometry readout presents extremely high counting precision and the assay’s coefficient of variation can be as low as 0.5%. sRCA-flow cytometry readout can be applied to detect the tumor mutations down to 1/100,000 in the circulating tumor cell-free DNA.In paper II, we applied the super RCA method into the in situ sequencing protocol to enhance the amplified mRNA detection tags for better signal-to-noise ratios. The sRCA products co-localize with primary RCA products generated from the gene specific padlock probes and remain as a single individual object in during the sequencing step. The enhanced sRCA products is 100% brighter than regular RCA products and the detection efficiency at least doubled with preserved specificity using sRCA compared to standard RCA.In paper III, we described a highly specific and efficient molecular switch mechanism namely RCA reporter. The switch will initiate the rolling circle amplification only in the presence of correct target sequences. The RCA reporter mechanism can be applied to recognize single stranded DNA sequences, mRNA sequences and sequences embedded in the RCA products.In paper IV, we established the solid phase Proximity Ligation Assay against the SOX10 protein using poly clonal antibodies. Using this assay, we found elevated SOX10 in serum at high frequency among vitiligo and melanoma patients. While the healthy donors below the threshold.
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| 7. |
- Chen, Lei, 1985-, et al.
(författare)
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Rare Mutation Detection in Blood Plasma Using sRCA Molecule Counting Probes
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Annan publikation (övrigt vetenskapligt/konstnärligt)abstract
- Problems in biology and medicine frequently require the ability to observe, evaluate, andcount even extremely rare macromolecules in biological samples. In particular, rare tumorspecific mutations in plasma provide valuable insights in the course of malignant disease andresponses to therapy, but simpler assay techniques are needed. We describe herein a rapidand exquisitely specific means to recognize and magnify detection signals from individualmolecules to easily recorded levels via a process we call super rolling circle amplification(sRCA). We demonstrate the ability of this technique to enumerate tumor-specific sequencevariants in plasma from cancer patients via flow cytometry at very high efficiency, with specificityadequate to detect single nucleotide mutant sequences among 100,000 copies of thenormal sequence in a 3 hr protocol. And the mutation analysis data generated from patientctDNA samples with our sRCA method are in high accordance with the patients’ primary tumorsequencing data.
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| 8. |
- Chen, Mo, et al.
(författare)
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Study on Efficient Fused Deposition Modelling of Thermoplastic Polyurethane Inflatable Wall Features for Airtightness
- 2020
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Ingår i: Swedish Production Symposium 2020.
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Konferensbidrag (refereegranskat)abstract
- The thermoplastic polyurethane (TPU) material is an elastomer that canbe used for inflatable products. Fused deposition modelling (FDM) is a widelyused additive manufacturing process for TPU material due to the capability ofgenerating complex structures with low cost. However, TPU is soft and thusdifficult to be extruded as continuously and uniformly as hard materials such aspolylactide by FDM. Inappropriate extruder structure and speed settings can leadto filament buckling problem, resulting in poor material filling quality, longprinting time and low printing success rate. This paper aims at improving the FDMprinting efficiency of TPU inflatable products by adding lateral support to thefilament and finding out the appropriate speed ranges for different wall featuresand thicknesses. Firstly, a filament guide sheet is designed as being inserted intothe gap between the drive gears and the bottom frame of the gear chamber in orderto prevent the soft TPU filament from buckling. Secondly, inflatable product wallfeatures are classified into floors, roofs and sidewalls and experiment for findingthe relationship between printing speed and airtightness is carried out. In order toverify the proposed solution, wall features are printed and the material fillingsobtained under different printing speeds are compared by measuring theairtightness of the wall features. Results show that the proposed filament guidesheet mitigates filament buckling, and the speed range that meets the airtightnessrequirement can be found for various wall features and thicknesses. In summary,the sealing of the filament feeding channel between the drive gears and the nozzle,as well as the speed optimisation according to product features, are essential forthe efficient printing of TPU inflatable products.
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| 9. |
- Daniel, Quentin, et al.
(författare)
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Water Oxidation Initiated by In Situ Dimerization of the Molecular Ru(pdc) Catalyst
- 2018
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Ingår i: ACS Catalysis. - : AMER CHEMICAL SOC. - 2155-5435. ; 8:5, s. 4375-4382
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Tidskriftsartikel (refereegranskat)abstract
- The mononuclear ruthenium complex [Ru(pdc)L-3] (H(2)pdc = 2,6-pyridinedicarboxylic acid, L = N-heterocycles such as 4-picoline) has previously shown promising catalytic efficiency toward water oxidation, both in homogeneous solutions and anchored on electrode surfaces. However, the detailed water oxidation mechanism catalyzed by this type of complex has remained unclear. In order to deepen understanding of this type of catalyst, in the present study, [Ru(pdc)(py)(3)] (py = pyridine) has been synthesized, and the detailed catalytic mechanism has been studied by electrochemistry, UV-vis, NMR, MS, and X-ray crystallography. Interestingly, it was found that once having reached the Ru-IV state, this complex promptly formed a stable ruthenium dimer [Ru-III(pdc)(py)(2)-O-Ru-IV(pdc)(py)(2)](+). Further investigations suggested that the present dimer, after one pyridine ligand exchange with water to form [Ru-III(pdc)(py)(2)-O-Ru-IV(pdc)(py)(H2O)](+), was the true active species to catalyze water oxidation in homogeneous solutions.
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| 10. |
- Ikebuchi, Ryoyo, et al.
(författare)
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Human proteins incorporated into tick-borne encephalitis virus revealed by in situ proximity ligation
- 2020
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Ingår i: Biochemical and Biophysical Research Communications - BBRC. - : ACADEMIC PRESS INC ELSEVIER SCIENCE. - 0006-291X .- 1090-2104. ; 525:3, s. 714-719
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Tidskriftsartikel (refereegranskat)abstract
- Host proteins incorporated into virus particles have been reported to contribute to infectivity and tissue-tropism. This incorporation of host proteins is expected to be variable among viral particles, however, protein analysis at single-virus levels has been challenging. We have developed a method to detect host proteins incorporated on the surface of virions using the in situ proximity ligation assay (isPLA) with rolling circle amplification (RCA), employing oligonucleotide-conjugated antibody pairs. The technique allows highly selective and sensitive antibody-based detection of viral and host proteins on the surface of individual virions. We detected recombinant noninfectious sub-viral particles (SVPs) of tick-borne encephalitis virus (TBEV) immobilized in microtiter wells as fluorescent particles detected by regular fluorescence microscopy. Counting the particles in the images enabled us to estimate individual TBEVSVP counts in different samples. Using isPLA we detected individual calnexin-, CD9-, CD81-, CD29-and CD59-positive SVPs among the viral particles. Our data suggests that a diversity of host proteins may be incorporated into TEBV, illustrating that isPLA with digital counting enables single-virus analysis of host protein incorporation. (C) 2020 The Authors. Published by Elsevier Inc.
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| 11. |
- Ji, Qinglei, et al.
(författare)
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A Flexible 4D Printing Service Platform for Smart Manufacturing
- 2020
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Ingår i: Swedish Production Symposium.
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Konferensbidrag (refereegranskat)abstract
- With the extensive application of 3D printing (3DP) in smartmanufacturing, 4D printing (4DP), which enhances 3D printed objects with shapemorphing ability by using smart materials, has shown significant industrial potentialand attracted tremendous attention. One key concern of 4DP is how to effectivelyand quickly meet different production and application requirements considering thecomplexity of materials and diversity of stimulus methods. In order to provide ageneral research platform for 4DP researchers, a flexible 4DP service platform isproposed. Components and modules for building 4DP and test systems are modeledand virtualized to form the different resources. These resources are then integratedvirtually or physically to provide some basic functions such as a 3D displacementstage or a visual monitoring system. According to different 4DP requirements, thesefunctions are then encapsulated into services to serve different research. Theplatform enables a variety of 4DP applications in smart manufacturing environmentssuch as 4D printed magnetic medical robots, test platform for studying the 4DPresponse, etc. A case study on designing a ferromagnetic 4DP platform based on theservice platform is performed to prove the feasibility of the method.
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| 12. |
- Ji, Qinglei, et al.
(författare)
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Design and closed loop control of a 3D printed soft actuator
- 2020
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Ingår i: 2020 16th IEEE International Conference on Automation Science and Engineering (CASE). - : IEEE. ; , s. 842-848
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Konferensbidrag (refereegranskat)abstract
- Soft robots, made of soft materials such as di-electric elastomer or shape memory polymers, have receivedtremendous attentions due to its dexterousness, flexibility andsafety compared with rigid robots. However, wider applicationof soft robots is limited due to their complex fabrication processand poor controllability. Here, we introduce a closed loopcontrolled soft actuator that is fully 3D printed with flexiblematerial. The structure of the soft actuator is optimized withFinite Element Method (FEM) to acquire shortest fabricationtime and highest deformation for same stimulus input. A desk-top Fused Deposition Modeling (FDM) 3D printer is used forlow-cost fabrication of such actuators. A webcamera is used forthe image feedback which offers the real time shape monitoringof the soft actuator. An output feedback Proportional IntegralDerivative (PID) controller with lowpass filter is developed withpole placement design method based on a data-driven modelof the 3D printed soft actuator. The controller is implementedto regulate the input air pressure to ensure a fast-response, precise and robust shape changing for any work environments.
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| 13. |
- Ji, Qinglei, et al.
(författare)
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Feedback control for the precise shape morphing of 4D printed shape memory polymer
- 2021
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Ingår i: IEEE Transactions on Industrial Electronics. - : IEEE. - 0278-0046 .- 1557-9948. ; , s. 12698-12707
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Tidskriftsartikel (refereegranskat)abstract
- Four-dimensional printing (4DP) is a newly emerged technology that uses smart materials for additive manufacturing and thus enables shape and/or property change upon stimulus after the printing process. Present study on 4DP has been focused on open loop stimulus, which can hardly ensure high shape precision and predictable final states. In this paper, a new closed loop 4DP (CL4DP) process supplementing 4D printed actuation with closed loop control methods is proposed. Image feedback is used for enhancing the conventional open loop 4DP morphing process and a controller is implemented to regulate the intensity of the stimulus accordingly in real-time. To achieve precise control, a nonlinear affine system model is built by model identification with measurement data to describe the dynamic shape recovery process of the 4D printed Shape Memory Polymer (SMP). Precise shape control is achieved and the effects of controller parameters on the precision of CL4DP are studied. Traditionally, SMP has a discrete number of selected steady states. With CL4DP, such steady states can be continuous and arbitrary.
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| 14. |
- Ji, Qinglei, 1993-
(författare)
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Learning-based Control for 4D Printing and Soft Robotics
- 2022
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Exploiting novel sensors and actuators made of flexible and smart materials becomes a new trend in robotics research. The studies on the design, production, and control of the new type of robots motivate the research fields of soft robots and 4D printed robots. 3D Printing (3DP) is an additive manufacturing technology that is widely used in printing flexible materials to fabricate soft robots. 4D Printing (4DP) combines 3DP technologies with smart materials to produce transformable devices. 4DP first prints structures with specifically designed responsive materials. When external stimuli such as temperature, voltage, or magnetic field are applied to the printed structure, it changes shape in a programmable way. The shape morphing property of 4DP makes it a novel approach to the actuators of robots.The employment of these special materials empowers these new robots with better compliance and adaptability to the working environment. However, compared with the rigid counterparts, they also have complex dynamic properties such as substantial non-linearity and time-variance. These factors make the precise modeling and robust control of these new robots challenging and thus hinder their potential applications. Focusing on soft robotic systems enabled by 3DP and 4DP approaches, this dissertation studies both traditional and Machine Learning (ML)-based approaches to the modeling, perception, and control of soft, non-linear, and time-variant robotic systems. The main contributions of this dissertation are:The scheme of Closed-Loop (CL) controlled 4DP (CL4DP) using temperature stimulated Shape Memory Polymer (SMP) is designed and validated numerically and experimentally. The feedback control system increases the precision and robustness of the shape morphing process of 4D printed SMP. Applications of CL4DP are explored.Data-driven model identification methods are applied to learn the dynamic model of the shape morphing process of CL4DP and the learned model has good quality to support model-based control design. Model-free and adaptive Reinforcement Learning (RL) controllers are developed to deal with the non-linearity and time variance of 4D printed actuators. To improve the stability and quick adaptability, a concise basis function set is selected instead of blindly using Deep Neural Networks (DNNs).A quadruped robot enabled by soft actuators and its simulation model are developed. The computation efficiency and model accuracy of the simulator are studied and optimized by comparing different simulation methods such as Finite Element Method (FEM) and lumped parameter method.The optimal walking gait pattern of a soft-legged quadruped robot is found by grid parameter search and RL with a physics based simulation model. To speed up the RL training process, modeling tricks are used to reduce the simulation time of the model and curriculum learning is used to reduce the learning time.A soft sensor made by printable conductive materials and 3DP is designed and optimally calibrated to estimate the shape of a pneumatically driven soft actuator. The geometry of the soft sensor is optimally designed for the best linearity, hysteresis and drift properties. The online estimation is based on a linear regression model learned from experimental data.A pneumatically driven soft gripper is developed by 3DP, the printable soft sensor, and pole-placement control methods. The operation of the gripper does not require an external image feedback system to measure its shape, which is estimated by the integrated soft sensor. The position feedback by the soft sensor and the controller by the pole-placement method enable the soft gripper to perform complex tasks with high precision.
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| 15. |
- Ji, Qinglei, et al.
(författare)
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Optimal shape morphing control of 4D printed shape memory polymer based on reinforcement learning
- 2022
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Ingår i: Robotics and Computer-Integrated Manufacturing. - : Elsevier. - 0736-5845 .- 1879-2537. ; 73
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Tidskriftsartikel (refereegranskat)abstract
- 4D printing technology, as a new generation of Additive Manufacturing methods, enables printed objectsto further change their shapes or other properties upon external stimuli. One main category of 4D printingresearch is 4D printed thermal Shape Memory Polymer (SMP). Its morphing process has large time delay, isnonlinear time variant, and susceptible to unpredictable disturbances. Reaching an arbitrary position with highprecision is an active research question. This paper applies the Reinforcement Learning (RL) method to developan optimal control method to perform closed loop control of the SMP actuation. Precise and prompt shapemorphing is achieved compared with previous control methods using a PI controller. The training efforts of RLare further reduced by simplifying the optimal control policy using the structural property of the prior trainedresults. Customized protective visors against COVID-19 are fabricated using the proposed control method.
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| 16. |
- Li, Fusheng, 1985-, et al.
(författare)
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Control the O-O bond formation pathways by immobilizing molecular catalysts on glassy carbon via electrochemical polymerization
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Annan publikation (övrigt vetenskapligt/konstnärligt)abstract
- Molecular water oxidation catalysts Ru-bda (1) and Ru-pda (2) are electrochemically polymerized on glassy carbon (GC) electrodes. Reaction orders and kinetic isotope effects (KIE) of the corresponding electrodes are studied. Results indicate that poly-1@GC goes through a radical coupling pathway. By adding poly-styrene (PSt) as a “blocking unit” in the poly-1, the radical coupling process of Ru-bda is blocked, and poly-1+PSt@GC catalyzes water oxidation through the water nucleophilic attack pathway. In comparison, catalyst 2, which oxidizes water via water nucleophilic attack path in homogeneous systems, goes through a radical coupling pathway as well when 2 is polymerized on glassy carbon (poly-2@GC).
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| 17. |
- Li, Shiyu, et al.
(författare)
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Nanoarrays on Passivated Aluminum Surface for Site-Specific Immobilization of Biomolecules
- 2018
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Ingår i: ACS Applied Bio Materials. - : American Chemical Society (ACS). - 2576-6422. ; 1:1, s. 125-135
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Tidskriftsartikel (refereegranskat)abstract
- The rapid development of biosensing platforms for highly sensitive and specific detection raises the desire of precise localization of biomolecules onto various material surfaces. Aluminum has been strategically employed in the biosensor system due to its compatibility with CMOS technology and its optical and electrical properties such as prominent propagation of surface plasmons. Herein, we present an adaptable method for preparation of carbon nanoarrays on aluminum surface passivated with poly(vinylphosphonic acid) (PVPA). The carbon nanoarrays were defined by means of electron beam induced deposition (EBID) and they were employed to realize site-specific immobilization of target biomolecules. To demonstrate the concept, selective streptavidin/neutravidin immobilization on the carbon nanoarrays was achieved through protein physisorption with a significantly high contrast of the carbon domains over the surrounding PVPA-modified aluminum surface. By adjusting the fabrication parameters, local protein densities could be varied on similarly sized nanodomains in a parallel process. Moreover, localization of single 40 nm biotinylated beads was achieved by loading them on the neutravidin-decorated nanoarrays. As a further demonstration, DNA polymerase with a streptavidin tag was bound to the biotin-beads that were immobilized on the nanoarrays and in situ rolling circle amplification (RCA) was subsequently performed. The observation of organized DNA arrays synthesized by RCA verified the nanoscale localization of the enzyme with retained biological activity. Hence, the presented approach could provide a flexible and universal avenue to precise localizing various biomolecules on aluminum surface for potential biosensor and bioelectronic applications.
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| 18. |
- Robelius, Anna, 1977-, et al.
(författare)
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Managing leukemia patients via liquid biopsy and super rolling circle amplification (superRCA)
- 2023
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Ingår i: Journal of Internal Medicine. - : John Wiley & Sons. - 0954-6820 .- 1365-2796. ; 294:2, s. 228-237
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Tidskriftsartikel (refereegranskat)abstract
- The rapidly increasing availability of sequence information for tumor patients, combined with expanding treatment options, motivates efforts to monitor the course of disease for individual patients by analyzing patient-specific mutations in liquid biopsies, as highly specific markers of the malignancy. We discuss the suitability of established molecular methods to monitor patients with malignancies, in particular leukemias, comparing these to the recently developed super rolling circle amplification technique for highly sensitive, parallel measurements of mutant sequences using readily available instruments. The very high sensitivity for tumor-specific mutations-in combination with low cost and ready access at clinics-promises to allow routine monitoring of increasing numbers of tumor patients, in order to initiate improved treatments at the earliest timepoint possible, when necessary. A method with high-enough accuracy to enable monitoring in peripheral blood rather than bone marrow samples would present a great practical advantage, not least from the patient perspective. We describe scenarios in which sufficiently sensitive, inexpensive methods for mutational analysis can provide valuable guidance for the clinician in choosing among therapeutic options and adjusting ongoing treatment and help to promptly identify recurrences of disease in treated patients.
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| 19. |
- Sandberg, Emma, et al.
(författare)
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Sensitive and Specific Analyses of Colorectal Cancer Recurrence through Multiplex superRCA Mutation Detection in Blood Plasma
- 2024
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Ingår i: Cancers. - : MDPI. - 2072-6694. ; 16:3
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Tidskriftsartikel (refereegranskat)abstract
- Mutation analysis of circulating tumor DNA (ctDNA) has applications in monitoring of colorectal cancer (CRC) patients for recurrence. Considering the low tumor fraction of ctDNA in cell-free DNA (cfDNA) isolated from blood plasma, the sensitivity of the detection method is important. Here, plasma DNA collected at diagnosis and follow-up from 25 CRC patients was analyzed using a multiplex superRCA mutation detection assay. The assay was also performed on genomic DNA (gDNA) from tumor and normal tissue from 20 of these patients. The lower limit of detection for most sequence variants was in the range of 10−5, while when analyzing cfDNA from plasma with a typical input of 33 ng, the practical detection limit was ~10−4 or 0.01% mutant allele frequency (MAF). In 17 of 19 patients with identified hotspot mutations in tumor gDNA, at least one hotspot mutation could be detected in plasma DNA at the time of diagnosis. The MAF increased at subsequent time points in four of the patients who experienced a clinical relapse. Multiplex superRCA analysis of the remaining six patients did not reveal any hotspot mutations. In conclusion, multiplex superRCA assays proved suitable for monitoring CRC patients by analyzing hotspot mutations in cfDNA, and dynamic changes in MAF were observed in patients with clinical relapse.
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| 20. |
- Vaskaninova, Valeria, et al.
(författare)
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Marginal dentition and multiple dermal jawbones as the ancestral condition of jawed vertebrates
- 2020
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Ingår i: Science. - : AMER ASSOC ADVANCEMENT SCIENCE. - 0036-8075 .- 1095-9203. ; 369:6500, s. 211-
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Tidskriftsartikel (refereegranskat)abstract
- The dentitions of extant fishes and land vertebrates vary in both pattern and type of tooth replacement. It has been argued that the common ancestral condition likely resembles the nonmarginal, radially arranged tooth files of arthrodires, an early group of armoured fishes. We used synchrotron microtomography to describe the fossil dentitions of so-called acanthothoracids, the most phylogenetically basal jawed vertebrates with teeth, belonging to the genera Radotina, Kosoraspis, and Tlamaspis (from the Early Devonian of the Czech Republic). Their dentitions differ fundamentally from those of arthrodires; they are marginal, carried by a cheekbone or a series of short dermal bones along the jaw edges, and teeth are added lingually as is the case in many chondrichthyans (cartilaginous fishes) and osteichthyans (bony fishes and tetrapods). We propose these characteristics as ancestral for all jawed vertebrates.
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| 21. |
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| 22. |
- Wu, Chenglin, et al.
(författare)
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Profiling and genotyping individual mRNA molecules through in situ sequencing of super rolling circle amplification products
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Annan publikation (övrigt vetenskapligt/konstnärligt)abstract
- We have recently developed a technology for localized sequence library preparation with rolling-circle amplification (RCA) as an approach for in situ sequencing. This method involves generation of clonally amplified and specially confined substrates for next-generation sequencing within the preserved context of cells and tissues. Our approach combines padlock probing, RCA, and sequencing-by-ligation chemistry that can resolve expression profiles of sets of genes and mutations in tissues without loss of histological context. Like other fluorescence-based assays, it can be hindered by high level of background fluorescence. To achieve high signal-to-noise ratios, we now describe a method to boost the amplification generated by RCA of padlock probes in situ by super RCA (sRCA). In this technique, a second padlock probe is hybridized, ligated and amplified on the first RCA product for enhanced, localized amplification. We describe and compare different sRCA strategies where gap-fill ligation was showed to be most efficient. The sRCA products co-localize and have comparable sizes as RCA products but they display at least two fold higher signal intensity. This increase in signal to noise also proved to result in two folds increase in the number of sRCA products detected. By combining sRCA with in situ sequencing for highly multiplex detection in tissue a four-time increase was seen. In summary, we demonstrate that sRCA can significantly increase the performance of padlock-based in situ sequencing for gene expression profiling of tissue sections, enabling detection of low abundant transcripts and the analysis of also highly auto-fluorescent samples.
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| 23. |
- Xie, Yuan, et al.
(författare)
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LGR5 promotes tumorigenicity and invasion of glioblastoma stem-like cells and is a potential therapeutic target for a subset of glioblastoma patients
- 2019
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Ingår i: Journal of Pathology. - : John Wiley & Sons. - 0022-3417 .- 1096-9896. ; 247:2, s. 228-240
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Tidskriftsartikel (refereegranskat)abstract
- Glioblastoma (GBM) is the most common and lethal primary malignant brain tumor which lacks efficient treatment and predictive biomarkers. Expression of the epithelial stem cell marker Leucine-rich repeat-containing G-protein coupled receptor 5 (LGR5) has been described in GBM, but its functional role has not been conclusively elucidated. Here, we have investigated the role of LGR5 in a large repository of patient-derived GBM stem cell (GSC) cultures. The consequences of LGR5 overexpression or depletion have been analyzed using in vitro and in vivo methods, which showed that, among those with highest LGR5 expression (LGR5(high)), there were two phenotypically distinct groups: one that was dependent on LGR5 for its malignant properties and another that was unaffected by changes in LGR5 expression. The LGR5-responding cultures could be identified by their significantly higher self-renewal capacity as measured by extreme limiting dilution assay (ELDA), and these LGR5(high)-ELDA(high) cultures were also significantly more malignant and invasive compared to the LGR5(high)-ELDA(low) cultures. This showed that LGR5 expression alone would not be a strict marker of LGR5 responsiveness. In a search for additional biomarkers, we identified LPAR4, CCND2, and OLIG2 that were significantly upregulated in LGR5-responsive GSC cultures, and we found that OLIG2 together with LGR5 were predictive of GSC radiation and drug response. Overall, we show that LGR5 regulates the malignant phenotype in a subset of patient-derived GSC cultures, which supports its potential as a predictive GBM biomarker. Copyright (c) 2018 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.
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