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  1. Article ; Online: DNA Cross-Reactivity of the CDC-Specified SARS-CoV-2 Specimen Control Leads to Potential for False Negatives and Underreporting of Viral Infection.

    Rosebrock, Adam P

    Clinical chemistry

    2020  Volume 67, Issue 2, Page(s) 435–437

    MeSH term(s) COVID-19/diagnosis ; COVID-19/epidemiology ; COVID-19/virology ; COVID-19 Testing ; Centers for Disease Control and Prevention, U.S. ; Cross Reactions ; DNA, Viral/chemistry ; False Negative Reactions ; Humans ; Pandemics ; Polymerase Chain Reaction/methods ; SARS-CoV-2/genetics ; United States
    Chemical Substances DNA, Viral
    Language English
    Publishing date 2020-11-12
    Publishing country England
    Document type Letter ; Research Support, N.I.H., Extramural
    ZDB-ID 80102-1
    ISSN 1530-8561 ; 0009-9147
    ISSN (online) 1530-8561
    ISSN 0009-9147
    DOI 10.1093/clinchem/hvaa284
    Database MEDical Literature Analysis and Retrieval System OnLINE

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  2. Article ; Online: Patient DNA cross-reactivity of CDC SARS-nCoV2 extraction control leads to potential false negative results

    Rosebrock, Adam P

    bioRxiv

    Abstract: Detecting RNA viruses such as SARS-nCoV2 requires careful handling of inherently labile RNA. Molecular tests incorporate controls to verify the presence of intact RNA and ensure that all steps of an assay have succeeded. The Centers for Disease Control ... ...

    Abstract Detecting RNA viruses such as SARS-nCoV2 requires careful handling of inherently labile RNA. Molecular tests incorporate controls to verify the presence of intact RNA and ensure that all steps of an assay have succeeded. The Centers for Disease Control and Prevention (CDC)-specified extraction control recognizes both reverse-transcribed RNA and human genomic DNA. Human DNA co-purified from nasopharyngeal swabs by multiple clinically-used RNA extraction approaches, is sufficient for a strong "extraction control positive" signal using the CDC design, creating the potential for false-negative results. Moreover, DNA cross-reactivity precludes control of RNA integrity in real-world samples. A properly designed control that monitors sample collection, extraction, reverse transcription, and qPCR processes is essential and demonstrated here. This control can be immediately implemented in the CDC testing workflow.
    Keywords covid19
    Language English
    Publishing date 2020-05-14
    Publisher Cold Spring Harbor Laboratory
    Document type Article ; Online
    DOI 10.1101/2020.05.13.094839
    Database COVID19

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  3. Article ; Online: Patient DNA cross-reactivity of the CDC SARS-CoV-2 extraction control leads to an inherent potential for false negative results

    Rosebrock, Adam P.

    bioRxiv

    Abstract: Testing for RNA viruses such as SARS-CoV-2 requires careful handling of inherently labile RNA during sample collection, clinical processing, and molecular analysis. Tests must include fail-safe controls that affirmatively report the presence of intact ... ...

    Abstract Testing for RNA viruses such as SARS-CoV-2 requires careful handling of inherently labile RNA during sample collection, clinical processing, and molecular analysis. Tests must include fail-safe controls that affirmatively report the presence of intact RNA and demonstrate success of all steps of the assay. A result of “no virus signal” is insufficient for clinical interpretation: controls must also say “The reaction worked as intended and would have found virus if present.” Unfortunately, a widely used test specified by the US Centers for Disease Control and Prevention (CDC) incorporates a control that does not perform as intended and claimed. Detecting SARS-CoV-2 with this assay requires both intact RNA and successful reverse transcription. The CDC-specified control does not require either of these, due to its inability to differentiate human genomic DNA from reverse-transcribed RNA. Patient DNA is copurified from nasopharyngeal swabs during clinically-approved RNA extraction and is sufficient to return an “extraction control success” signal using the CDC design. As such, this assay fails-unsafe: truly positive patient samples return a false-negative result of “no virus detected, control succeeded” following any of several readily-encountered mishaps. This problem affects tens-of-millions of patients worth of shipped assays, but many of these flawed reagents have not yet been used. There is an opportunity to improve this important diagnostic tool. As demonstrated here, a re-designed transcript-specific control correctly monitors sample collection, extraction, reverse transcription, and qPCR detection. This approach can be rapidly implemented and will help reduce truly positive patients from being incorrectly given the all-clear. One Sentence Summary A widely-used COVID-19 diagnostic is mis-designed and generates false-negative results, dangerously confusing “No” with “Don’t know” – but it’s fixable
    Keywords covid19
    Publisher BioRxiv; WHO
    Document type Article ; Online
    DOI 10.1101/2020.05.13.094839
    Database COVID19

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  4. Article ; Online: Targeted full-scan LC-MS metabolomics: simultaneous quantitation of knowns and feature discovery provide the best of both worlds.

    Rosebrock, Adam P

    Bioanalysis

    2017  Volume 9, Issue 1, Page(s) 5–8

    MeSH term(s) Animals ; Chromatography, Liquid/methods ; Humans ; Mass Spectrometry/methods ; Metabolome ; Metabolomics/methods
    Language English
    Publishing date 2017-01
    Publishing country England
    Document type Journal Article
    ISSN 1757-6199
    ISSN (online) 1757-6199
    DOI 10.4155/bio-2016-0256
    Database MEDical Literature Analysis and Retrieval System OnLINE

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  5. Article ; Online: Analysis of the Budding Yeast Cell Cycle by Flow Cytometry.

    Rosebrock, Adam P

    Cold Spring Harbor protocols

    2017  Volume 2017, Issue 1

    Abstract: DNA synthesis is one of the landmark events in the cell cycle: ... ...

    Abstract DNA synthesis is one of the landmark events in the cell cycle: G
    MeSH term(s) Cell Cycle ; Flow Cytometry/methods ; Fluorescent Dyes/metabolism ; Saccharomyces cerevisiae/cytology ; Saccharomyces cerevisiae/physiology ; Staining and Labeling/methods
    Chemical Substances Fluorescent Dyes
    Language English
    Publishing date 2017--03
    Publishing country United States
    Document type Journal Article
    ISSN 1559-6095
    ISSN (online) 1559-6095
    DOI 10.1101/pdb.prot088740
    Database MEDical Literature Analysis and Retrieval System OnLINE

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  6. Article ; Online: Synchronization of Budding Yeast by Centrifugal Elutriation.

    Rosebrock, Adam P

    Cold Spring Harbor protocols

    2017  Volume 2017, Issue 1

    Abstract: In yeast, cell size is normally tightly linked to cell cycle progression. Centrifugal elutriation is a method that fractionates cells based on the physical properties of cell size-fluid drag and buoyant density. Using a specially modified centrifuge and ... ...

    Abstract In yeast, cell size is normally tightly linked to cell cycle progression. Centrifugal elutriation is a method that fractionates cells based on the physical properties of cell size-fluid drag and buoyant density. Using a specially modified centrifuge and rotor system, cells can be physically separated into one or more cohorts of similar size and therefore cell cycle position. Small G
    MeSH term(s) Cell Division ; Centrifugation/methods ; Microbiological Techniques/methods ; Saccharomycetales/growth & development ; Saccharomycetales/physiology
    Language English
    Publishing date 2017--03
    Publishing country United States
    Document type Journal Article
    ISSN 1559-6095
    ISSN (online) 1559-6095
    DOI 10.1101/pdb.prot088732
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  7. Article ; Online: Methods for Synchronization and Analysis of the Budding Yeast Cell Cycle.

    Rosebrock, Adam P

    Cold Spring Harbor protocols

    2017  Volume 2017, Issue 1

    Abstract: Like other eukaryotes, budding yeast temporally separate cell growth and division. DNA synthesis is distinct from chromosome segregation. Storage carbohydrates are accumulated slowly and then rapidly liquidated once per cycle. Cyclin-dependent kinase ... ...

    Abstract Like other eukaryotes, budding yeast temporally separate cell growth and division. DNA synthesis is distinct from chromosome segregation. Storage carbohydrates are accumulated slowly and then rapidly liquidated once per cycle. Cyclin-dependent kinase associates with multiple different transcriptionally and posttranslationally regulated cyclins to drive the cell cycle. These and other crucial events of cellular growth and division are limited to narrow windows of the cell cycle. Many experiments in the yeast laboratory treat a culture of cells as a homogeneous mixture. Measurements of asynchronous cultures are, however, confounded by the presence of cells in various cell cycle stages; measuring a population average in unsynchronized cells provides at best a decreased signal and at worst an artifactual result. A number of experimentally tractable methods have been developed to generate populations of yeast cells that are synchronized with respect to cell cycle phase. Robust methods for determining cell cycle position have also been developed. These methods are introduced here.
    MeSH term(s) Cell Cycle ; Microbiological Techniques/methods ; Saccharomycetales/growth & development ; Saccharomycetales/physiology
    Language English
    Publishing date 2017--03
    Publishing country United States
    Document type Journal Article
    ISSN 1559-6095
    ISSN (online) 1559-6095
    DOI 10.1101/pdb.top080630
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  8. Article ; Online: Synchronization and Arrest of the Budding Yeast Cell Cycle Using Chemical and Genetic Methods.

    Rosebrock, Adam P

    Cold Spring Harbor protocols

    2017  Volume 2017, Issue 1

    Abstract: The cell cycle of budding yeast can be arrested at specific positions by different genetic and chemical methods. These arrests enable study of cell cycle phase-specific phenotypes that would be missed during examination of asynchronous cultures. Some ... ...

    Abstract The cell cycle of budding yeast can be arrested at specific positions by different genetic and chemical methods. These arrests enable study of cell cycle phase-specific phenotypes that would be missed during examination of asynchronous cultures. Some methods for arrest are reversible, with kinetics that enable release of cells back into a synchronous cycling state. Benefits of chemical and genetic methods include scalability across a large range of culture sizes from a few milliliters to many liters, ease of execution, the absence of specific equipment requirements, and synchronization and release of the entire culture. Of note, cell growth and division are decoupled during arrest and block-release experiments. Cells will continue transcription, translation, and accumulation of protein while arrested. If allowed to reenter the cell cycle, cells will do so as a population of mixed, larger-than-normal cells. Despite this important caveat, many aspects of budding yeast physiology are accessible using these simple chemical and genetic tools. Described here are methods for the block and release of cells in G
    MeSH term(s) Cell Cycle Checkpoints/drug effects ; Cell Cycle Checkpoints/radiation effects ; Cell Cycle Proteins/genetics ; Cell Division/drug effects ; Cell Division/radiation effects ; Genetics, Microbial/methods ; Hot Temperature ; Mating Factor/metabolism ; Microbiological Techniques/methods ; Mutant Proteins/metabolism ; Saccharomycetales/drug effects ; Saccharomycetales/growth & development ; Saccharomycetales/physiology ; Saccharomycetales/radiation effects
    Chemical Substances Cell Cycle Proteins ; Mutant Proteins ; Mating Factor (61194-02-3)
    Language English
    Publishing date 2017--03
    Publishing country United States
    Document type Journal Article
    ISSN 1559-6095
    ISSN (online) 1559-6095
    DOI 10.1101/pdb.prot088724
    Database MEDical Literature Analysis and Retrieval System OnLINE

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  9. Article: MOB rules: Antibiotic Exposure Reprograms Metabolism to Mobilize

    Liu, Yongjin / LaBonte, Sandra / Brake, Courtney / LaFayette, Carol / Rosebrock, Adam P / Caudy, Amy A / Straight, Paul D

    bioRxiv : the preprint server for biology

    2024  

    Abstract: Antibiotics have dose-dependent effects on exposed bacteria. The medicinal use of antibiotics relies on their growth-inhibitory activities at sufficient concentrations. At subinhibitory concentrations, exposure effects vary widely among different ... ...

    Abstract Antibiotics have dose-dependent effects on exposed bacteria. The medicinal use of antibiotics relies on their growth-inhibitory activities at sufficient concentrations. At subinhibitory concentrations, exposure effects vary widely among different antibiotics and bacteria.
    Language English
    Publishing date 2024-03-20
    Publishing country United States
    Document type Preprint
    DOI 10.1101/2024.03.20.585991
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  10. Article ; Online: Metabolite Extraction from

    Rosebrock, Adam P / Caudy, Amy A

    Cold Spring Harbor protocols

    2017  Volume 2017, Issue 9, Page(s) pdb.prot089086

    Abstract: Prior to mass spectrometric analysis, cellular small molecules must be extracted and separated from interfering components such as salts and culture medium. To ensure minimal perturbation of metabolism, yeast cells grown in liquid culture are rapidly ... ...

    Abstract Prior to mass spectrometric analysis, cellular small molecules must be extracted and separated from interfering components such as salts and culture medium. To ensure minimal perturbation of metabolism, yeast cells grown in liquid culture are rapidly harvested by filtration as described here. Simultaneous quenching of metabolism and extraction is afforded by immediate immersion in low-temperature organic solvent. Samples prepared using this method are suitable for a range of downstream liquid chromatography-mass spectrometry analyses and are stable in solvent for >1 yr at -80°C.
    Language English
    Publishing date 2017-09-01
    Publishing country United States
    Document type Journal Article
    ISSN 1559-6095
    ISSN (online) 1559-6095
    DOI 10.1101/pdb.prot089086
    Database MEDical Literature Analysis and Retrieval System OnLINE

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