Protein detection using proximity-dependent DNA ligation assays

Simon, Fabrice; Gullberg, Mats; Jarvius, Jonas; Olsson, Charlotta; Pietras, Kristian; Gústafsdóttir, Sigrún M.; Östman, Arne; Landegren, Ulf

doi:10.1038/nbt0502-473

Cited by 1,292 publications

(1,143 citation statements)

References 23 publications

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“…This recently developed tool combines an antibody-based approach with QPCR (Fredriksson et al, 2002;Gullberg et al, 2004). Using an oligonucleotide-labeled polyclonal antibody (or two different monoclonal antibodies specific for different epitopes on the same protein), one batch of 'left primer' antibody and one batch of 'right primer' antibody are biotinylated and coupled to specific 'left' and 'right' oligonucleotides; when the left and right antibodies are colocalized on the target protein, the left and right oligos may then hybridize with a linking oligo and be amplified using PCR.…”

Section: Western Blot Analysesmentioning

confidence: 99%

Postmortem Brain: An Underutilized Substrate for Studying Severe Mental Illness

McCullumsmith

Hammond

Shan

et al. 2013

Neuropsychopharmacol

106

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We propose that postmortem tissue is an underutilized substrate that may be used to translate genetic and/or preclinical studies, particularly for neuropsychiatric illnesses with complex etiologies. Postmortem brain tissues from subjects with schizophrenia have been extensively studied, and thus serve as a useful vehicle for illustrating the challenges associated with this biological substrate. Schizophrenia is likely caused by a combination of genetic risk and environmental factors that combine to create a disease phenotype that is typically not apparent until late adolescence. The complexity of this illness creates challenges for hypothesis testing aimed at understanding the pathophysiology of the illness, as postmortem brain tissues collected from individuals with schizophrenia reflect neuroplastic changes from a lifetime of severe mental illness, as well as treatment with antipsychotic medications. While there are significant challenges with studying postmortem brain, such as the postmortem interval, it confers a translational element that is difficult to recapitulate in animal models. On the other hand, data derived from animal models typically provide specific mechanistic and behavioral measures that cannot be generated using human subjects. Convergence of these two approaches has led to important insights for understanding molecular deficits and their causes in this illness. In this review, we discuss the problem of schizophrenia, review the common challenges related to postmortem studies, discuss the application of biochemical approaches to this substrate, and present examples of postmortem schizophrenia studies that illustrate the role of the postmortem approach for generating important new leads for understanding the pathophysiology of severe mental illness.

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Section: Western Blot Analysesmentioning

confidence: 99%

Postmortem Brain: An Underutilized Substrate for Studying Severe Mental Illness

McCullumsmith

Hammond

Shan

et al. 2013

Neuropsychopharmacol

106

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“…As soon as we gain the capacity to register single protein molecules and their complexes, we shall be able to address the problem of compiling proteomic maps in any biological material on a single-molecule level. The alternative approach in this research area, which will probably find wide application in the future, lies in the development of methods enabling replication of protein molecules [27] or the use of PCR for monitoring the antigen/antibody interaction reactions (immuno-PCR [28], immuno-RCA [29]), and the method for proximal coupling of aptamers [30]). However, insufficient development of these methods does not yet allow us to consider them as a basis for highly productive proteomic investigations.…”

Section: Using the 2-d-lc-ms/ms Method We Could Not Identifymentioning

confidence: 99%

AFM fishing nanotechnology is the way to reverse the Avogadro number in proteomics

et al. 2007

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Future development of proteomics may be hindered by limitations in the concentration sensitivity of widespread technological approaches. The concentration sensitivity limit (CSL) of currently used approaches, like 2-DE/LC separation coupled with MS detection, etc., varies from 10 29 to 10 212 M. Therefore, proteomic technologies enable detection of up to 20% of the protein species present in the plasma. New technologies, like atomic force microscopy (AFM molecular detector), enable the counting of single molecules, whereas biospecific fishing can be used to capture these molecules from the biomaterial. At the same time, fishing also has thermodynamic limitations due to the reversibility of the binding. In cases where the fishing becomes irreversible, its combination with an AFM detector enables the registration of single protein molecules, and that opens up a way to lower the CSL down to the reverse Avogadro number.

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“…[26][27][28] Hematoxylin and eosin-stained tissue sections were analyzed to identify a tumor-rich region that was used to create a tissue microarray. For sample pretreatment, a 4-l-thick tissue microarray section was deparaffinized and epitope-retrieved in citrate buffer, pH 6.0 (DBS, Pleasanton, Calif) under pressurized boiling for 5 minutes.…”

Section: Ag-11mentioning

confidence: 99%

A phase 2 trial of single‐agent bevacizumab given in an every‐3‐week schedule for patients with recurrent high‐grade gliomas

Raizer

Grimm

Chamberlain

et al. 2010

Cancer

139

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BACKGROUND:The authors evaluated a 3-week schedule of bevacizumab in patients with recurrent high-grade glioma (HGG). METHODS: Patients received bevacizumab 15 mg/kg every 3 weeks and were evaluated every 6 weeks until tumor progression. Tissue correlates were used to quantify tumor content of vascular endothelial growth factor A (VEGFA) and vascular endothelial growth factor receptor-2 (VEGFR2). RESULTS: Of 61 patients who were treated (35 men and 26 women; median age, 52 years; age range, 21-78 years), 50 patients had glioblastoma multiforme (GBM), and 11 patients had anaplastic glioma (AG). The median number of previous chemotherapies was 2 (range, 1-5 previous chemotherapies), and 16 patients had received !3 previous chemotherapies. The median number of bevacizumab doses was 4 (range, 1-20 doses), and 45% of patients received >5 doses. The toxicities observed were primarily grade 1 and 2, and the most common were fatigue, hypertension, and headache. One grade 2 intratumoral bleed and 1 bowel perforation were reported. For patients with GBM, the 6-month progression-free survival rate was 25%, the median time to tumor progression was 10.8 weeks, and the median overall survival was 25.6 weeks. The best response included a partial response in 15 patients (24.5%) and stable disease in 31 patients (50.8%) patients; radiographic recurrence patterns included increased changes in fluid attenuation inversion recovery (24%) and multifocal recurrence (20%). The median survival after bevacizumab failure was 10 weeks. The ratio of tumor VEGFA/VEGFR2 was increased in patients aged >55 years; an increased VEGFA/VEGFR2 ratio was correlated nonsignificantly with decreased survival (P ¼ .052). CONCLUSIONS: An every-3-week schedule of bevacizumab had antitumor activity and was relatively nontoxic for patients with recurrent HGG. The predictive value of VEGFA/VEGFR2 in tumor will require validation in a larger patient cohort. Cancer 2010;116:5297-305.

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Protein detection using proximity-dependent DNA ligation assays

Cited by 1,292 publications

References 23 publications

Postmortem Brain: An Underutilized Substrate for Studying Severe Mental Illness

Postmortem Brain: An Underutilized Substrate for Studying Severe Mental Illness

AFM fishing nanotechnology is the way to reverse the Avogadro number in proteomics

A phase 2 trial of single‐agent bevacizumab given in an every‐3‐week schedule for patients with recurrent high‐grade gliomas

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