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Fourth Annual Digital PCR - 第4回デジタルPCR年次学会 -
2015年11月3 - 4日
米国、カリフォルニア州、サンディエゴ

デジタルPCR学会:精密診断のための技術およびツール– 第1日



ポリメラーゼ連鎖反応(PCR)は核酸の検出と定量評価にとって非常に重要なツールです。診断試験所における不可欠なものとして、PCRは科学的ニーズとともに発展をつづけ、最新のデジタルPCR(dPCR)となっています。dPCRは感度と特異性が向上しただけでなく、多くの場合、処理能力も向上しました。Cambridge Healthtech Instituteによる第4回デジタルPCR年次学会では、臨床診断のための重要なPCRの進化に焦点を当て続けていきます。例年通り、技術プロバイダーや早期導入者、新規プラットフォーム参入者が集結し、交流を深めながらデジタルPCRの能力と限界、将来的応用について議論します。今年は、dPCRに関する技術に焦点を当てるだけでなく、dPCRとNGSとの間のインターフェース、新興技術および研究分野にも注目します。その他、処理困難な試料のためのソリューション、試料処理能力を向上させるためのソリューションをはじめ、細胞外DNAの研究、デジタル検出のためのガイドラインとベストプラクティスなども扱います。

プログラムアドバイザー

Jim Huggett, B.Sc. (Hons), Ph.D., Science Leader, Nucleic Acid Metrology, Molecular & Cell Biology, LGC
N. Somanath Bhat, Ph.D., Acting Senior Research Scientist, Bioanalysis Group, Department of Industry and Science, National Measurement Institute, Australia


主な議題

  • 技術的配慮事項:チップ、液滴から多重化まで
  • DPCRを用いたバリデーションおよび参照基準
  • デジタルPCRデータ解析
  • dPCRとNGSのインターフェース
  • 絶対的定量
  • コピー数の変動
  • 少数標的検出

主な講演者


Jim HuggettJim Huggett, BSc (Hons), Ph.D., Science Leader, Nucleic Acid Metrology, Molecular & Cell Biology, LGC GroupFilip

 

Filip JankuFilip Janku, M.D., Ph.D., Assistant Professor, Investigational Cancer Therapeutics (Phase I Program), MD Anderson Cancer Center

 

Valerie TalyValerie Taly, Ph.D., Group Leader, CNRS Researcher, UMR S1147, University of Paris Descartes, CNRS

 

ショートコース*

デジタルPCR:技術的概要 

Jim Huggett, BSc (Hons), Ph.D., Science Leader, Nucleic Acid Metrology, Molecular & Cell Biology, LGC Group

単一細胞解析技術:準備状況と能力 

Michael Masterman-Smith, Ph.D., CEO, Harmony Biosciences, Inc.


*別途登録が必要です。



11月3日(火)


第1日:デジタルPCRの技術的側面

7:30 am 参加登録とモーニングコーヒー

8:25 議長による開会の挨拶

N. Somanath Bhat, Ph.D., Acting Senior Research Scientist, Bioanalysis Group, Department of Industry and Science, National Measurement Institute 

 

技術的配慮事項:チップ、液滴から多重化まで

8:30 結晶デジタルPCRの導入

Dangla RemiRémi Dangla, Ph.D., President and Co-Founder, Stilla Technologies

Stilla Technologies unveils its unique tool for high precision genetic analysis: Crystal digital PCR. Taking advantage of groundbreaking microfluidic advances, Crystal digital PCR relies on a single consumable to perform on-chip PCR in monolayer droplet arrays. This innovative approach increases multiplexing capacities in digital PCR and is readily amenable to large-scale, high-throughput clinical studies, thanks to a simple and fast workflow.

9:00 液滴デジタルMDAを用いた全ゲノム増幅

Minsoung RheeMinsoung Rhee, Ph.D., Postdoctoral Research Fellow, Biological Science and Technology, Sandia National Labs

We demonstrated for the first time that whole genome amplification by MDA can be performed in picoliter emulsion droplets, resulting in improved performance over corresponding reactions carried out at conventional microliter scale. De novo assembly of E. coli genome sequences from our droplet MDA and conventional tube MDA has revealed that droplet MDA showed more uniform coverage for all length of the genome and ~>99% of specific amplification.

9:30 統合された包括的液滴デジタル検出を用いた血中希少バイオマーカーの即時検出

Dong-Ku KangDong-Ku Kang, Ph.D., Assistant Research Scientist, Sue and Bill Gross Stem Cell Research Center, Pharmaceutical Sciences & Biomedical Engineering, University of California, Irvine; Co-Founder and CSO, VeloxBiosystems

Rapid and sensitive diagnosis remains a major unmet challenge in food industry and medical applications. In this presentation, I will discuss a new technology called "Integrated Comprehensive Droplet Digital Detection Technology" (IC 3D) that is able to rapidly (1-2 h) and selectively detect rare pathogens/or rare biomarkers from milliliters (mLs) of complex media (e.g., unprocessed whole blood) at single-cell sensitivity in a one-step, homogenous, and culture-free reaction. This platform technology also has the potential to introduce a new paradigm in rapid detection for various diseases including cancer (targeting circulating tumor cells), neurological disorder (e.g., Alzheimer's disease), and infectious diseases (e.g., Lyme disease, HIV, and Ebola).

10:00 休憩と展示会およびポスター発表の見学


DPCRを用いたバリデーションおよび参照基準

10:45 DNA標準物質のためのデジタルPCR

Somanath BhatN. Somanath Bhat, Ph.D., Acting Senior Research Scientist, Bioanalysis Group, Department of Industry and Science, National Measurement Institute

Accurate, reliable and reproducible quantification of nucleic acids is vital for many diagnostic applications and in routine laboratory testing. Digital PCR has the potential to not only improve quantitative nucleic acid analysis, but also to be considered as a reference method for certification of nucleic acid reference materials (RMs). This presentation will discuss how this technology was applied to characterize DNA RMs and discuss factors affecting reliability of the results.

11:15 臨床評価におけるdPCRによる計数の潜在的役割:KRAS SNPの例

Jim HuggettJim Huggett, BSc (Hons), Ph.D., Science Leader, Nucleic Acid Metrology, Molecular & Cell Biology, LGC

Unlike qPCR, dPCR does not require a calibration curve for quantitative analysis as the partitioning required to perform the technique enables DNA to be directly counted, or enumerated. This characteristic is fairly unique and opens a number of possibilities when considering clinical measurement, either through direct measurement using dPCR or in its use to support other methods, like qPCR, through the quantification of reference materials. This talk will discuss the work of the European Metrology Research Programme funded project Bio SITrace (http://biositrace.lgcgroup.com/) which is investigating the accuracy of dPCR when measuring rare single nucleotide polymorphisms in cell free DNA.

11:45 スポンサー後援プレゼンテーション

12:15 pm ランチョンプレゼンテーションまたは各自での昼食

1:00 セッションの中断


デジタルPCRデータ解析

1:25 議長による挨拶

Jim Huggett, BSc (Hons), Ph.D., Science Leader, Nucleic Acid Metrology, Molecular & Cell Biology, LGC 

 

1:30 一般化された線形混合モデルによる液滴デジタルPCRのデータ解析

Oliver ThasOliver Thas, Ph.D., Professor, Biostatistics, Ghent University (Belgium) and University of Wollongong (Australia)

Target quantification with ddPCR depends heavily on the Poisson assumption. We demonstrate how Generalized Linear Mixed Models (GLMM) can be used for the data-analysis. GLMM is very flexible and allows for analyzing many designs, including dealing with replicates, run or plate effects and one or more references. The method can be used for absolute quantification, CNV and gene expression, and it allows for standard error and confidence interval calculation and hypothesis testing.

2:00 HHistoMosaicの、マイクロ流体マトリクスにおけるin situ PCRによるCRC組織内のG12V KRAS検出

Emil KartalovEmil Kartalov, Ph.D., Assistant Professor, Pathology, Keck School of Medicine, University of Southern California

We present the experimental proof of principle of HistoMosaic - a novel technique for in situ genetic analysis of cancer tissue sections. HistoMosaic offers a high-throughput, low-cost, high-sensitivity means of detection and localization of rare mutations conferring drug resistance to cancer tissue, while the morphological information is preserved and co-registered with the genetic information. This ability would allow proper stratification of cancer patients, so the right drug is given to the right patient. HistoMosaic also has wide applicability in fundamental research and drug development.

2:30 スポンサー後援プレゼンテーション

3:00 休憩と展示会およびポスター発表の見学


NGSとDPCRとの間のインターフェース

3:30 NGSおよびデジタルPCRによる進行性腫瘍からの複数生検タイプにおける変異検出

Errin L. LagowErrin L. Lagow, Ph.D., Senior Scientific Liaison, Asuragen, Inc.

Targeted NGS permits detection of low-frequency somatic mutations in tumor biopsies, but call confidence may require independent assessment. Multiple biopsy types from advanced tumors were analyzed with the Quantidex™ PanCancer Panel, designed, developed, and cGMP-manufactured by Asuragen, and with digital PCR. Low frequency mutations were identified in FFPE tissue and confirmed in matched frozen tissue. Digital PCR demonstrated high utility for resolving discordant calls for samples with low frequency mutations.

4:00 コピー数の変動:デジタルPCR、NanoStringおよび次世代シーケンシング

Reinhold PollnerReinhold Pollner, Ph.D., Director, Clinical Trial Assay Development, Genoptix, Inc., a Novartis company

A variety of different technologies can be utilized to determine copy number variations in clinical samples. My presentation will focus on how three digital technologies - digital PCR, NanoString and Next-Generation Sequencing are used to determine copy number variations in clinical trial samples for patient stratification or exploratory purposes.

4:30 デジタルPCRによるNGSライブラリの正確な定量評価のための課題と戦略

Peter SchweitzerPeter Schweitzer, Ph.D., Director, Genomics Facility, Institute of Biotechnology, Cornell University

One critical step in producing high quality DNA sequence data in a timely and cost efficient manner is accurately quantifying Illumina sequencing libraries. Digital PCR (dPCR) offers several advantages over real-time PCR. One significant challenge is accurately performing the large dilution required and I'll describe an internal reference developed to correct for variability during dilution. I'll also describe our experiences with dPCR assays for Illumina libraries using several different platforms.

5:00 ウェルカムレセプションと展示会およびポスター発表の見学

6:00 第1日の閉会


* 不測の事態により、事前の予告なしにプログラムが変更される場合があります。



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