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  • 1. Alksnis, M.
    et al.
    Lindberg, A Michael
    Department of Medical Genetics, Uppsala University.
    Stålhanske, POK
    Hultberg, H.
    Pettersson, U.
    Use of synthetic oligodeoxyribonucleotides for type-specific identification of coxsackie B viruses1989In: Molecular and Cellular Probes, ISSN 1044-7431, E-ISSN 1095-9327, Vol. 3, no 2, p. 103-108Article in journal (Refereed)
    Abstract [en]

    Synthetic oligodeoxyribonucleotides were used for type-specific identification of members of the coxsackie B virus group by nucleic acid hybridization. Two pairs of oligonucleotide chains were constructed based on nucleotide sequences in the VP1 regions of coxsackieviruses B3 and B4. Each labelled probe had a length of 24 nucleotides. The results showed that the oligonucleotide hybridized in a type-specific manner when assayed with extracts from cells infected with all different coxsackie B viruses. A method based on similar principles may thus be used for enterovirus typing.

  • 2.
    Andrae, Johanna
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology.
    Hansson, Inga
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology.
    Afink, G B
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology.
    Nistér, Monica
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology.
    Platelet-derived growth factor receptor-alpha in ventricular zone cells and in developing neurons.2001In: Molecular and Cellular Probes, ISSN 1044-7431, E-ISSN 1095-9327, Vol. 17, no 6Article in journal (Refereed)
    Abstract [en]

    Cells in the early neuroepithelium differentiate and give rise to all cells in the central nervous system (CNS). The ways from a multipotent CNS stem cell to specialized neurons and glia are not fully understood. Using immunohistochemistry we found that neuroepithelial cells express the platelet-derived growth factor receptor-alpha (PDGFR-alpha) in the neural plate at embryonic day 8.5 and onwards in the neural tube. The protein was polarized to ventricular endfeet. Furthermore, PDGFR-alpha expression was localized to cells undergoing early neuronal development. We also found PDGFR-alpha expression in developing granule cells in the postnatal cerebellum, in Purkinje cells in the adult cerebellum and on processes of developing dorsal root ganglion cells. Previous reports mainly describe PDGFR-alpha expression in oligodendrocyte precursors and glial cells. We believe, in line with a few previous reports, that the PDGFR-alpha in addition marks a pool of undifferentiated cells, which are able to differentiate into neurons.

  • 3.
    Ansell - Schultz, Anna
    et al.
    Linköping University, Department of Clinical and Experimental Medicine, Divison of Neurobiology. Linköping University, Faculty of Medicine and Health Sciences.
    Reyes, Juan
    Linköping University, Department of Clinical and Experimental Medicine, Divison of Neurobiology. Linköping University, Faculty of Medicine and Health Sciences.
    Samuelsson, My
    Linköping University, Department of Clinical and Experimental Medicine, Divison of Neurobiology. Linköping University, Faculty of Medicine and Health Sciences. Region Östergötland, Center for Diagnostics, Clinical pathology.
    Hallbeck, Martin
    Linköping University, Department of Clinical and Experimental Medicine, Divison of Neurobiology. Linköping University, Faculty of Medicine and Health Sciences. Region Östergötland, Center for Diagnostics, Clinical pathology.
    Reduced retromer function results in the accumulation of amyloid-beta oligomers2018In: Molecular and Cellular Probes, ISSN 1044-7431, E-ISSN 1095-9327, Vol. 93, p. 18-26Article in journal (Refereed)
    Abstract [en]

    Alzheimers disease (AD) is a neurodegenerative disorder characterized by a progressive loss of multiple cognitive functions. Accumulation of amyloid beta oligomers (oA beta) play a major role in the neurotoxicity associated with the disease process. One of the early affected brain regions is the hippocampus, wherein a reduction of the vacuolar protein sorting-associated protein 35 (VPS35), the core protein comprising the retromer complex involved in cellular cargo sorting, has been identified. To investigate the role of the retromer function on the accumulation and clearance of oA beta, we reduced retromer function by selectively inhibiting VPS35 gene expression using siRNA in differentiated neuronal SH-SY5Y cells. As cell-to-cell transfer of oA beta to new brain regions is believed to be important for disease progression we investigated the effect of VPS35 reduction both in cells with direct uptake of oA beta and in cells receiving oA beta from donor cells. We demonstrate that reduced retromer function increases oA beta accumulation in both cell systems, both the number of cells containing intracellular oA beta and the amount within them. This effect was shown at different time points and regardless if the AD originated from the extracellular milieu or via a direct neuronal cell-to-cell transfer. Interestingly, not only did reduced VPS35 cause oA beta accumulation, but oA beta treatment alone also lead to a reduction of VPS35 protein content. The accumulated oA beta seems to co-localize with VPS35 and early endosome markers. Together, these findings provide evidence that reduced retromer function decreases the ability for neurons to transport and clear neurotoxic oA beta received through different routes resulting in the accumulation of oA beta. Thus, enhancing retromer function may be a potential therapeutic strategy to slow down the pathophysiology associated with the progression of AD.

  • 4. Bäckman, A
    et al.
    Lantz, P-G
    Rådström, P-G
    Olcén, Per
    Linköping University, Faculty of Health Sciences. Linköping University, Department of health and environment.
    Evaluation of an extended diagnostic PCR assay for detection and verification of the common causes of bacterial meningitis in CSF and other biological samples. 1999In: Molecular and Cellular Probes, ISSN 1044-7431, E-ISSN 1095-9327, Vol. 13, p. 49-60Article in journal (Refereed)
  • 5. Dahlén, P
    et al.
    Syvänen, Ann-Christine
    Hurskainen, P
    Kwiatkowski, M
    Sund, C
    Ylikoski, J
    Söderlund, H
    Lövgren, T
    Sensitive detection of genes by sandwich hybridization and time-resolved fluorometry1987In: Molecular and Cellular Probes, ISSN 1044-7431, E-ISSN 1095-9327, Vol. 1, no 2, p. 159-168Article in journal (Refereed)
    Abstract [en]

    Europium has been used as a non-radioactive marker in immunoassays as this metal can be detected with high sensitivity by time-resolved fluorometry. In this work streptavidin labeled with europium was used to detect biotinylated probes in a sandwich nucleic-acid hybridization assay with microtitration strips as the solid phase. pBR 322 plasmids were detected with a sensitivity of 4 × 105 molecules. As the sample is added in solution in sandwich hybridization, fast and simple sample pre-treatment can be used without encountering background problems. The method was applied to test bacterial samples of uropathogenic Escherichia coli strains for the presence of the β-lactamase gene.

  • 6. Gharizadeh, B.
    et al.
    Ohlin, A.
    Molling, P.
    Backman, A.
    Amini, B.
    Olcen, P.
    Nyrén, Pål
    KTH, Superseded Departments, Biochemistry and Biotechnology.
    Multiple group-specific sequencing primers for reliable and rapid DNA sequencing2003In: Molecular and Cellular Probes, ISSN 1044-7431, E-ISSN 1095-9327, Vol. 17, no 4, p. 203-210Article in journal (Refereed)
    Abstract [en]

    Pyrosequencing(TM) technology is a bioluminometric DNA sequencing method that employs a cascade of four enzymes to deliver sequence signals. To date this technology has been limited to the sequencing of short stretches of DNA. As an improvement to this technique, we have introduced a bacterial group-specific, multiple sequencing primer approach that circumvents sequencing of less informative semi-conservative regions of the 16S rRNA gene. This new approach is suitable for challenging templates, improving sequence data quality, avoiding sequencing of non-specific amplification products, lessening sequencing time, and moreover, this strategy should open the way for many new applications in the future. The group-specific, multiple sequencing primers can be applied in the Sanger dideoxy sequencing method as well. In addition, we have improved the chemistry of the Pyrosequencing system enabling sequencing of longer stretches of DNA, which allows numerous new applications.

  • 7.
    Gharizadeh, Baback
    et al.
    Department of Biotechnology, Stockholm Center for Physics, Astronomy and Biotechnology, Royal Institute of Technology (KTH), Stockholm, Sweden.
    Ohlin, Andreas
    Department of Pediatrics, Örebro University Hospital, Örebro, Sweden.
    Mölling, Paula
    Department of Clinical Microbiology, Örebro University Hospital, Sweden.
    Bäckman, Anders
    Department of Clinical Microbiology, Örebro University Hospital, Sweden.
    Amini, Bahram
    Department of Biotechnology, Stockholm Center for Physics, Astronomy and Biotechnology, Royal Institute of Technology (KTH), Stockholm, Sweden.
    Olcén, Per
    Department of Clinical Microbiology, Örebro University Hospital, Sweden.
    Nyrén, Pål
    Department of Biotechnology, Stockholm Center for Physics, Astronomy and Biotechnology, Royal Institute of Technology (KTH), Stockholm, Sweden.
    Multiple group-specific sequencing primers for reliable and rapid DNA sequencing2003In: Molecular and Cellular Probes, ISSN 1044-7431, E-ISSN 1095-9327, Vol. 17, no 4, p. 203-210Article in journal (Refereed)
    Abstract [en]

    Pyrosequencing™ technology is a bioluminometric DNA sequencing method that employs a cascade of four enzymes to deliver sequence signals. To date this technology has been limited to the sequencing of short stretches of DNA. As an improvement to this technique, we have introduced a bacterial group-specific, multiple sequencing primer approach that circumvents sequencing of less informative semi-conservative regions of the 16S rRNA gene. This new approach is suitable for challenging templates, improving sequence data quality, avoiding sequencing of non-specific amplification products, lessening sequencing time, and moreover, this strategy should open the way for many new applications in the future. The group-specific, multiple sequencing primers can be applied in the Sanger dideoxy sequencing method as well. In addition, we have improved the chemistry of the Pyrosequencing system enabling sequencing of longer stretches of DNA, which allows numerous new applications.

  • 8.
    Gharizadeh, Baback
    et al.
    Stanford Univ, Stanford Genome Technol Ctr.
    Zheng, Biying
    Karolinska Inst, Dept Mol Med.
    Akhras, Michael
    KTH, School of Biotechnology (BIO).
    Ghaderi, Mehran
    Karolinska Univ Hosp, Clinical Pathology/Cytology.
    Jejelowo, Olufisayo
    Texas So Univ.
    Strander, Björn
    Gothenburg Univ, Sahlgrens Acad, Ctr Oncol.
    Nyrén, Pål
    KTH, School of Biotechnology (BIO).
    Wallin, Keng-Ling
    Karolinska Inst, Dept Mol Med.
    Pourmand, Nader
    Stanford Univ, Stanford Genome Technol Ctr.
    Sentinel-base DNA genotyping using multiple sequencing primers for high-risk human papillomaviruses2006In: Molecular and Cellular Probes, ISSN 1044-7431, E-ISSN 1095-9327, Vol. 20, no 3-4, p. 230-238Article in journal (Refereed)
    Abstract [en]

    Despite the various technologies in place for genotyping human papillomaviruses (HPV), clinical use and clinical research demand a method that is fast, more reliable and cost-effective. The technology described here represents a breakthrough development in that direction. By combining the method of multiple sequencing primers with DNA sequencing, we have developed a rapid assay for genotyping HPV that relies on the identification of a single, type-specific 'sentinel' base. As described here, the prototype assay has been developed to recognize the 12 most high-risk HPV types (HPV-16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58 and 59) and is capable of recognizing and simultaneously genotyping multiple HPV co-infections. By providing sequence information on multiple HPV infections, this method eliminates the need for labor- and cost-intensive PCR cloning. These proof-of-concept studies establish the assay to be accurate, reliable, rapid, flexible, and cost-effective, providing evidence of the feasibility this technique for use in clinical settings.

  • 9. Heurling, Kerstin
    et al.
    Ashton, Nicholas J
    Leuzy, Antoine
    Zimmer, Eduardo R
    Blennow, Kaj
    Zetterberg, Henrik
    Eriksson, Jonas
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Preclinical PET-MRI Platform. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Preparative Medicinal Chemistry.
    Lubberink, Mark
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Surgical Sciences, Radiology.
    Schöll, Michael
    Synaptic vesicle protein 2A as a potential biomarker in synaptopathies2019In: Molecular and Cellular Probes, ISSN 1044-7431, E-ISSN 1095-9327, Vol. 97, p. 34-42Article in journal (Refereed)
    Abstract [en]

    Measuring synaptic density in vivo using positron emission tomography (PET) imaging-based biomarkers targeting the synaptic vesicle protein 2A (SV2A) has received much attention recently due to its potential research and clinical applications in synaptopathies, including neurodegenerative and psychiatric diseases. Fluid-based biomarkers in proteinopathies have previously been suggested to provide information on pathology and disease status that is complementary to PET-based measures, and the same can be hypothesized with respect to SV2A. This review provides an overview of the current state of SV2A PET imaging as a biomarker of synaptic density, the potential role of fluid-based biomarkers for SV2A, and related future perspectives.

  • 10. Jungell-Nortamo, A
    et al.
    Syvänen, Ann-Christine
    Luoma, P
    Söderlund, H
    Nucleic acid sandwich hybridization: enhanced reaction rate with magnetic microparticles as carriers1988In: Molecular and Cellular Probes, ISSN 1044-7431, E-ISSN 1095-9327, Vol. 2, no 4, p. 281-288Article in journal (Refereed)
    Abstract [en]

    A method for the detection of nucleic acid hybrids using the sandwich hybridization technique with magnetic polystyrene microparticles as the solid support is described. The capture DNA is coupled to the polystyrene-hydroxy surface of the particles through p-toluenesulfonyl chloride activation. The use of microparticles results in a substantial increase in the reaction rate compared to filter hybridization, without decreasing the sensitivity of detection. Polyethylene glycol additionally enhances the reaction rate. The use of magnetic microparticles allows rapid and convenient collection of the formed hybrids.

  • 11.
    Lundberg, Gunilla A.
    Örebro University, Department of Clinical Medicine.
    A rapid one-tube PCR method for simultaneously differentiating homozygotes and heterozygotes of the Sp1 binding site polymorphism in collagen type I alpha 12007In: Molecular and Cellular Probes, ISSN 1044-7431, E-ISSN 1095-9327, Vol. 21, no 3, p. 239-241Article in journal (Refereed)
    Abstract [en]

    Rapid detection of single-base changes is fundamental to molecular medicine. PCR amplification of specific alleles (PASA) has previously been used as a rapid method of genotyping single-nucleotide changes, but one reaction is required for each allele. This paper describes a Bidirectional PASA (Bi-PASA) method, which was developed to distinguish between homozygotes and heterozygotes in one PCR reaction. The method is tested using the Sp1 polymorphism in Collagen type la I. The results demonstrate that Bi-PASA is a simple and rapid method for detecting the zygosity of the polymorphism in a single PCR reaction. (c) 2006 Elsevier Ltd. All rights reserved.

  • 12. Parkkinen, S
    et al.
    Mäntyjärvi, R
    Syrjänen, K
    Syvänen, Ann-Christine
    Ranki, M
    Sandwich hybridization in solution: a rapid method to screen HPV 16 DNA in cervical scrapes1989In: Molecular and Cellular Probes, ISSN 1044-7431, E-ISSN 1095-9327, Vol. 3, no 1, p. 1-11Article in journal (Refereed)
    Abstract [en]

    A solution hybridization method is introduced as a rapid diagnostic method for demonstration of papillomavirus DNA in cervical scrapes. 32P-Labelled detector probe and the biotinylated capture probes were hybridized with DNA of the specimen after pretreatment by boiling in alkaline SDS. After 4 h of hybridization the hybrids were collected onto avidin coated beads and measured. The sensitivity of the method was 1–5 × 105 HPV 16 DNA molecules. Cervical carcinoma cell lines CaSki and SiHa were informative as to the sensitivity of the solution hybridization and the in situ hybridization methods. CaSki cells containing about 700 HPV 16 DNA copies per cell were positive by both methods. SiHa cells with one HPV 16 DNA copy per cell were positive by the sandwich assay but remained negative in the in situ test. A series of 126 cervical scrapes collected from consecutive patients participating in a follow-up study for cervical HPV infection were tested for HPV 16 DNA by both methods. The detection rate of the sandwich test was 19/126 (15%) and that of the in situ method 21/126 (17%) yielding 26 diagnoses altogether. Twelve of these were obtained by one method only. The results obtained by studying the cervical cell lines and repeated specimens taken from constantly HPV 16 positive patients suggested that the two methods can measure different types of infections and thus complement each other in the diagnosis of cervical HPV infections.

  • 13. Ruettger, Anke
    et al.
    Feige, Jens
    Slickers, Peter
    Schubert, Evelyn
    Morre, Servaas A.
    Pannekoek, Yvonne
    Herrmann, Björn
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Clinical Bacteriology.
    de Vries, Henry J. C.
    Ehricht, Ralf
    Sachse, Konrad
    Genotyping of Chlamydia trachomatis strains from culture and clinical samples using an ompA-based DNA microarray assay2011In: Molecular and Cellular Probes, ISSN 1044-7431, E-ISSN 1095-9327, Vol. 25, no 1, p. 19-27Article in journal (Refereed)
    Abstract [en]

    Current typing methods of Chlamydia (C) trachomatis are mainly based on the diversity of the ompA gene, which is coding for the major outer membrane protein A. The present study aimed at facilitating genotyping of strains of this obligate intracellular human pathogen by developing a DNA microarray assay using the ArrayTube (TM) format for individual samples and the ArrayStrip (TM) format for higher throughput. The new test is exploiting multiple discriminatory sites by involving a total of 61 oligonucleotide probes representing genotype-specific polymorphisms in variable domains 1, 2 and 4 of the ompA gene. After multiplex amplification of these domains using biotinylated primers, the sample is hybridized in the microarray vessel under highly stringent conditions. The resulting binding pattern is genotype specific, thus allowing direct identification. We were able to show that DNA from each of the currently accepted genotypes (serovars) yielded a unique, theoretically expected and distinct hybridization pattern. The assay was also shown to be highly sensitive as a dilution containing the equivalent of 1 inclusion-forming unit was still correctly genotyped. In addition, when 62 clinical samples were examined and compared to PCR-RFLP typing results, the genotype was correctly identified by the DNA microarray in all cases. The present test is easy to handle and economically affordable, and it allows genotyping of C. trachomatis to be accomplished within a working day, thus lending itself for epidemiological studies and routine diagnosis.

  • 14.
    Tärnberg, Maria
    et al.
    Linköping University, Department of Clinical and Experimental Medicine, Clinical Microbiology. Linköping University, Faculty of Health Sciences.
    Nilsson, Lennart E.
    Linköping University, Faculty of Health Sciences. Linköping University, Department of Clinical and Experimental Medicine, Clinical Microbiology.
    Monstein, Hans-Jürg
    Linköping University, Department of Clinical and Experimental Medicine, Clinical Microbiology. Linköping University, Faculty of Health Sciences.
    Molecular identification of blaSHV, blaLEN and blaOKP β-lactamase genes in Klebsiella pneumoniae by bi-directional sequencing of universal SP6- and T7-sequence-tagged blaSHV-PCR amplicons2009In: Molecular and Cellular Probes, ISSN 1044-7431, E-ISSN 1095-9327, Vol. 23, no 3-4, p. 195-200Article in journal (Refereed)
    Abstract [en]

    Plasmid encoded blaSHV enzymes represent an important sub-group of class A β-lactamases causing an ESBL-phenotype which is increasingly found in Enterobacteriaceae including Klebsiella pneumoniae. The chromosomally encoded β-lactamase blaLEN and blaOKP enzymes, which so far only have been found in K. pneumoniae, do not hydrolyse extended-spectrum cephalosporins. In the present study, multiple displacement amplified DNA derived from 20 K. pneumoniae clinical isolates with a blaSHV-like genotype was used in a universal SHV PCR assay using SP6- (forward) and T7- (reverse) sequence-tagged primers. Identification and differentiation of blaSHV, blaLEN and blaOKP genes was obtained by bi-directional amplicon sequencing using SP6- and T7-specific primers. Three well characterised K. pneumoniae strains having a SHV-genotype were included in the study. The bi-directional amplicon sequencing, covering 800 bp (93%) of the blaSHV, blaLEN and blaOKP enzyme encoding sequences, allowed for an unequivocal discrimination of SHV, LEN and OKP genes. Moreover, sequencing revealed the presence of blaSHV allelic variants in six K. pneumoniae isolates in which the amplicons had to be cloned accordingly. Based on deduced amino-acid sequences, a dendrogram was constructed. Seventeen out of 20 K. pneumoniae isolates with an ESBL-phenotype formed a SHV-like cluster, two were LEN-like, and one isolate was OKP-like. The PCR-based molecular typing method described here enables a rapid, reliable and cost-effective identification and differentiation of blaSHV, blaOKP and blaLEN genes.

  • 15.
    Zeng, Qing-Yin
    et al.
    Umeå University, Faculty of Science and Technology, Molecular Biology (Faculty of Science and Technology).
    Wang, Xiao-Ru
    Blomquist, Göran
    Development of mitochondrial SSU rDNA-based oligonucleotide probes for specific detection of common airborne fungi2003In: Molecular and Cellular Probes, ISSN 1044-7431, E-ISSN 1095-9327, Vol. 17, no 6, p. 281-288Article in journal (Refereed)
    Abstract [en]

    In this study we sequenced partial mitochondrial small subunit rDNA from 32 fungal strains representing 31 species from 16 genera. Most of these species are common airborne fungi and pathogens that may cause various public health concerns. Sequence alignment showed several conserved and highly variable regions. The variable regions were deployed to design oligonucleotide probes for each fungal species. The specificity of the designed probes was first examined through homology search against GenBank database then further verified through hybridization experiments to 38 fungal strains. A total of 23 probes were verified as specific to 15 fungal species commonly detected in living and working environments. These new probes will have potential applications in clinical diagnosis and public health-related environmental monitoring.

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