BEAMing: single-molecule PCR on microparticles in water-in-oil emulsions

Diehl, Frank; Li, Meng; He, Yiping; Kinzler, Kenneth W.; Vogelstein, Bert; Dressman, Devin

doi:10.1038/nmeth898

Cited by 441 publications

(389 citation statements)

References 14 publications

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“…New amplicons of smaller size were designed for the DNA purified from paraffin-embedded samples. When sequencing chromatograms were difficult to interpret in the DNA purified from tumor samples, we reevaluated the mutation in question either by cloning the PCR product and sequencing individual clones or by performing a BEAMing assay (27,28). Additional, more detailed methods are described in SI Methods.…”

Section: Methodsmentioning

confidence: 99%

“…Mutations present in a smaller fraction can generally not be distinguished from the background in sequencing chromatograms. To determine whether the mutations were present in a smaller but still sizable fraction of the tumor cell population, we evaluated a subset of the DNA samples via BEAMing (beads, emulsions, amplifications, magnetics) assay (see SI Methods) (27,28). We performed 20 BEAMing assays in seven patients, focusing on those mutations that appeared to be present in late-stage lesions but not in an earlier-stage lesion of the same patient (e.g., present in metastasis but not in the advanced colorectal carcinoma).…”

Section: Quantification Of the Level Of Mutations In Dnamentioning

confidence: 99%

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Comparative lesion sequencing provides insights into tumor evolution

Jones

Chen

Parmigiani

et al. 2008

Proc. Natl. Acad. Sci. U.S.A.

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747

631

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We show that the times separating the birth of benign, invasive, and metastatic tumor cells can be determined by analysis of the mutations they have in common. When combined with prior clinical observations, these analyses suggest the following general conclusions about colorectal tumorigenesis: (i) It takes Ϸ17 years for a large benign tumor to evolve into an advanced cancer but <2 years for cells within that cancer to acquire the ability to metastasize; (ii) it requires few, if any, selective events to transform a highly invasive cancer cell into one with the capacity to metastasize; (iii) the process of cell culture ex vivo does not introduce new clonal mutations into colorectal tumor cell populations; and (iv) the rates at which point mutations develop in advanced cancers are similar to those of normal cells. These results have important implications for understanding human tumor pathogenesis, particularly those associated with metastasis.cancer genetics ͉ colorectal cancer ͉ metastasis ͉ stem cells

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Section: Methodsmentioning

confidence: 99%

Section: Quantification Of the Level Of Mutations In Dnamentioning

confidence: 99%

Comparative lesion sequencing provides insights into tumor evolution

Jones

Chen

Parmigiani

et al. 2008

Proc. Natl. Acad. Sci. U.S.A.

Self Cite

747

631

View full text Add to dashboard Cite

show abstract

“…Since the PCR-products are double-stranded, a strand separation needs to be performed to isolate the single-stranded aptamers. Both requirements can be fulfilled by the use of BEAMing (Beads, Emulsion, Amplification, Magnetic), a combination of emulsion-PCR and solid phase-coupling according to Diehl and Hünniger [18,19]. Briefly, an according to the requirements modified Forward-Primer (5 -aminated or 5 -biotinylated), a Reverse-Primer coupled to magnetic beads and the other PCR components (polymerase, buffer, dNTPs, template and water) were mixed.…”

Section: Beamingmentioning

confidence: 99%

New SPR-based methods for analysis of allergenic agents used in wine treatment

Wessels¹,

Paschke-Kratzin²

2016

BIO Web Conf.

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Abstract. The use of allergenic agents in wines needs to be monitored by both producers and supervisory authorities for the protection of sensitive individuals. Currently established as the gold standard is the ELISAtechnique which relies on the affinity and specificity of antibodies. Since antibodies are produced in animals and current legislative developments demand the reduction of the use of animals for scientific purposes, the biosensor technology could be interesting for the quantification of allergens. By the combination of a technical device, the surface plasmon resonance, and a specific antibody it was possible to develop an antibody-based analytical method which is capable to detect lysozyme in quantities below 0.25 ppm. Since the biosensor is reusable multiple times, this approach can contribute to reduce the antibody consumption and therefore the use of animals for analysis. Furthermore, first insights in the use of new molecular receptors, aptamers were gained.

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“…These biomarkers have been explored as dynamic tools for monitoring response to systemic therapies and addressing the challenges posed by therapeutic resistance and intratumor genetic heterogeneity. In this way, the sensitive "BEAMing technique" (beads, emulsification, amplification, and magnetics) has demonstrated its ability for detecting and quantifying driver somatic mutations in plasma for breast and colorectal cancers [33,34]. As an example of the BEAMing technique, PIK3CA mutational status in plasma ctDNA and tumor tissues was concordant when samples were obtained simultaneously; however, when years elapsed from removal of tumor and the blood draw, discordant results were seen in as many as 20% of patients, demonstrating that biomarkers may change over time and clonal evolution may be one of the reasons.…”

Section: Pilot Studies Of Circulating Tumor Dna To Measure Tumor Genomentioning

confidence: 99%

Pilot Studies for Personalized Cancer Medicine: Focusing on the Patient for Treatment Selection

Mattos-Arruda

Rodón

2013

The Oncologist

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Advances in diagnostics and targeted therapies during the past decade have changed how oncology is viewed. "Stratified medicine" has emerged from the accumulated evidence garnered from matching targeted therapies with tumor molecular aberrations. Concomitantly, current knowledge derived from large-scale, massively parallel sequencing technologies and global research initiatives such as the international 1000 Genomes Project, the Cancer Genome Atlas, the International Cancer Genome Consortium, and publicly available catalogs such as the Catalogue of Somatic Mutations in Cancer and Genomics of Drug Sensitivity in Cancer have illuminated the utility of understanding the molecular basis of cancer through genome analysis. In addition, multiple collaborative efforts are widening the possibility of universally personalizing cancer care. Although several key challenges of personalized cancer medicine (PCM) need to be addressed, some pilot studies are transforming the way we analyze tumor tissue molecular aberrations, design clinical trials, and measure treatment efficacy. Taken together, these pilot studies are paving the way for clinical trials that are designed to empirically test the concept of PCM. In this paper, we describe lessons learned from the first pilot initiatives of PCM and how this knowledge is being used to design novel clinical trials.

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BEAMing: single-molecule PCR on microparticles in water-in-oil emulsions

Cited by 441 publications

References 14 publications

Comparative lesion sequencing provides insights into tumor evolution

Comparative lesion sequencing provides insights into tumor evolution

New SPR-based methods for analysis of allergenic agents used in wine treatment

Pilot Studies for Personalized Cancer Medicine: Focusing on the Patient for Treatment Selection

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