Nikiforova: #ampeurope2018 PPA is % of known vars detected by NGS cp to orthogonal method. Worksheet: https://t.co/Ph28MLN7Ln
11:40am May 1st 2018 via Twitter Web Client
Nikiforova: #ampeurope2018 59 samples per mutation type: shown to reach statistical power. Est PPA and PPV - b/c we… https://t.co/g8a9levdy7
11:35am May 1st 2018 via Twitter Web Client
Nikiforova: #ampeurope2018 And with each type of mutation (SNV, Indels, CNV, SVs), and samples with most common mut… https://t.co/SCV0HNyLDY
11:34am May 1st 2018 via Twitter Web Client
Nikiforova: #ampeurope2018 Est min read depth of >250 reads/amplicon or target for 5% MAF. Validation: using exact sample type.
11:33am May 1st 2018 via Twitter Web Client
Nikiforova: #ampeurope2018 Labs were all over, Larry Jennings proposed statistics (in paper), established distribution of TP, FP
11:32am May 1st 2018 via Twitter Web Client
Nikiforova: #ampeurope2018 Need to understand how the platform works, ref materials and standardizing methods. Depth for LoD determined.
11:31am May 1st 2018 via Twitter Web Client
Nikiforova: #ampeurope2018 Choice of platform, optimization & familiarization phase. All perform well, advantages.… https://t.co/2NBWCeU
11:30am May 1st 2018 via Twitter Web Client
Nikiforova: #ampeurope2018 2 methods: amplification or hybrid capture. Can design a commercial panel, or design. Na… https://t.co/UHW8CMbHCY
11:29am May 1st 2018 via Twitter Web Client
Nikiforova: #ampeurope2018 Validation, it's long, will make highlights and observations. Majority of clin labs: targeted over WGS, WES, RNA
11:27am May 1st 2018 via Twitter Web Client
Nikiforova: #ampeurope2018 And a long process, 14-18 months. JMD Validation: https://t.co/siZw2tdgJn Reporting: https://t.co/G22oEoj6GC
11:25am May 1st 2018 via Twitter Web Client
Nikiforova: #ampeurope2018 2012: the clinical NGS era began. At that time, no guidelines existed. In '14: initiated… https://t.co/IqA9exyXEL
11:24am May 1st 2018 via Twitter Web Client
Marina Nikiforova (Univ Pittsburgh, USA): #ampeurope2018 Practice Guidelines for Validation, Interpretation and Rep… https://t.co/o0bw7QNtvu
11:22am May 1st 2018 via Twitter Web Client
Q: Any look at ncRNA at the sc level? Pugh: #ampeurope2018 A current 'huge blind spot'. Bulk is never polyA; for sc… https://t.co/TRoKXCDQ64
11:10am May 1st 2018 via Twitter Web Client
Q: Can CITE-seq be only surface proteins? Pugh: #ampeurope2018 Currently, yes only surface to-date.
11:04am May 1st 2018 via Twitter Web Client
Pugh: #ampeurope2018 Concl with the Human Cell Atlas, the Chan Zuckerberg intitiative. https://t.co/Ts13bnB0gh
11:00am May 1st 2018 via Twitter Web Client
Pugh: #ampeurope2018 Upcoming methods - DroNc-Seq '17 Nature Methods https://t.co/x8EJboai7I And CITE-Seq https://t.co/LOmMoxTtHD
Pugh: #ampeurope2018 Combining cancer, healthy cell data highlights shared and private cellular populations.
10:52am May 1st 2018 via Twitter Web Client
Pugh: #ampeurope2018 Sees tumor cells, BTSC line, TIL immune cells. Treated each chr as a pathway. Inferred Chr 7, Chr 10 gain/loss
10:47am May 1st 2018 via Twitter Web Client
Pugh: #ampeurope2018 Large, freq loss of chr6 at scWGS, unexplained. Secret sauce is dissociation of brain tumor tissue. Shows tSNE1/2 plots
10:46am May 1st 2018 via Twitter Web Client
Pugh: #ampeurope2018 Glioblastomas have self-renewing cancer stem cells. Shows scRNA core set of genes, then tracked to chr6.
10:44am May 1st 2018 via Twitter Web Client
Pugh: #ampeurope2018 In cancer: scRNA to understand early, monitor, and see mech of resp/resistance. Stand Up to C… https://t.co/o3izjXltRg
10:40am May 1st 2018 via Twitter Web Client
Pugh: #ampeurope2018 Gene sets, pathways. '17 https://t.co/LOmMoxTtHD But a lack of a reference standard set.
Pugh: #ampeurope2018 Review of challenges in scRNA analysis '15 Nat Rev Genetics https://t.co/x1z5y9TBad Many ways to readout PCA.
10:37am May 1st 2018 via Twitter Web Client
Pugh: #ampeurope2018 Full length can also find coding mutaitons. Baran-Gale ref compares methods. Shows fig: 3 exons vs end-reads.
10:35am May 1st 2018 via Twitter Web Client
Pugh: #ampeurope2018 Difference: transcript structure throughout mRNA in SmartSeq 2, only 10's of thousands. 10X does only end-reads.
10:34am May 1st 2018 via Twitter Web Client
Pugh: #ampeurope2018 SmartSeq2 https://t.co/sjwypWsLB2 Cheap. $2-4/cell https://t.co/lYl7UQTxWl
10:33am May 1st 2018 via Twitter Web Client
Pugh: #ampeurope2018 Rev on sc techniques in immunol https://t.co/STQmZGLeAE Talk only about 10X Genomics, SmartSeq2
10:32am May 1st 2018 via Twitter Web Client
Pugh: #ampeurope2018 Immune infiltrate - but very different expression profiles on a single-cell level. Multiple pa… https://t.co/K8CE3k4S5J
10:28am May 1st 2018 via Twitter Web Client
Pugh: #ampeurope2018 Estimates of immune and stromal cell content, scored from bulk GEx data. Yoshihara et al '13 https://t.co/aV6EsrlwUx
10:26am May 1st 2018 via Twitter Web Client
Pugh: #ampeurope2018 '13 fig https://t.co/X3qRbM2NYM worked on infrastructure w/fresh biopsies with INSPIRE program, Pembro resp
10:25am May 1st 2018 via Twitter Web Client
Pugh: #ampeurope2018 Directs a research core for single-cell seq; a bowls of M&Ms, different colors, innards. Tumor… https://t.co/gfReNQ
10:24am May 1st 2018 via Twitter Web Client
Trevor Pugh (Princess Margaret, Toronto) #ampeurope2018 Simultaneous Analysis of Cancer Clones and Immune Microenv… https://t.co/NolmExAOje
10:22am May 1st 2018 via Twitter Web Client
Q: See differences across platforms? Brudzewsky: #ampeu2018 Some didn't see them and adjusted pipelines.
10:15am May 1st 2018 via Twitter Web Client
Q: ArcherDx variability in results between fusion types? Brudzewsky: #ampeu2018 Percent of fusion reads detected -… https://t.co/dU4h2uaLor
10:13am May 1st 2018 via Twitter Web Client
Q: ctDNA Brudzewsky: #ampeu2018 Have ctDNA and CNV products as well.
10:12am May 1st 2018 via Twitter Web Client
Brudzewsky: #ampeu2018 Has a poster with Helen Fernandez of Columbia Univ (O17) about this work.
10:11am May 1st 2018 via Twitter Web Client
Brudzewsky: #ampeu2018 Knowing 'ground truth' enables monitoring performance over time. 'NGS assay validation less daunting'
Brudzewsky: #ampeu2018 Shows nice dPCR vs NGS LOD chart of fusion RNA quantitation. Shows 6x runs on ArcherDx FusionPlex, and reprod
10:08am May 1st 2018 via Twitter Web Client
Brudzewsky: #ampeu2018 16 fusions, copies/uL determined, evaluated on Illumina, QIAGEN, ArcherDx, TMO.
10:06am May 1st 2018 via Twitter Web Client
Brudzewsky: #ampeu2018 SeraCare's approach - biosynthetic constructs w/5' cap, polyA; introduced to human reference… https://t.co/cEs0J9Ghw2
10:05am May 1st 2018 via Twitter Web Client
Brudzewsky: #ampeu2018 Unique challenges: NTRK is a new target, 'absence of proof is not proof of absence'
10:04am May 1st 2018 via Twitter Web Client
Brudzewsky: #ampeu2018 reference range, min input amt, LoD, analytical spec, analytical sens. And by mutation type, many things to consider.
10:01am May 1st 2018 via Twitter Web Client
Brudzewsky: #ampeu2018 Starts with the complexity of NGS workflow. Somatic panels need to consider for validation:… https://t.co/pSjGOxZz4m
10:00am May 1st 2018 via Twitter Web Client
Dan Brudzewsky (SeraCare US): #ampeu2018 Use of Highly Multiplexed Reference Materials to Facilitate Validation of… https://t.co/LTeMMjZTu2
9:59am May 1st 2018 via Twitter Web Client
Q: Other new mutations introduced? Saddar: #ampeu2018 Footprint-free lines, using new advances.
9:54am May 1st 2018 via Twitter Web Client
Q: Role of CNV in the lines? Saddar: #ampeu2018 Diploid, and homozygous. They do have other lines with CNVs.
Saddar: #ampeu2018 Have also done FFPE, slides, scrolls, matched frozen pellets, gDNA. Have also done ctDNA.
9:53am May 1st 2018 via Twitter Web Client
Saddar: #ampeu2018 Via AmpliSeq Hotspot panel w/MiSeq, measured 1.74% for EGFR del, 0.97% for T790M, 0.76% for KRAS G13D
9:52am May 1st 2018 via Twitter Web Client
Saddar: #ampeu2018 Deposit was 200K (not 20K), shows 10% 'coverage' of cell, shows TMO QS 3D digital PCR at 10%, 5% and 1%
9:50am May 1st 2018 via Twitter Web Client
Saddar: #ampeu2018 Cell types mixed with EGFR T790M, KRAS G13D, EGFR E746-A750 del, deposit 20K cells/slide. Then dPCR/NGS
9:49am May 1st 2018 via Twitter Web Client
