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
Saddar: #ampeu2018 Homology-directed repair if a repair template available, where precise point mutations, indels or tags inserted.
9:46am May 1st 2018 via Twitter Web Client
Saddar: #ampeu2018 As cell lines, relevant, reprod and reliable source. Cas9-mediated DSB, using gRNA directing Cas9 to site.
9:45am May 1st 2018 via Twitter Web Client
Saddar: #ampeu2018 Need for FNA: most common for lung ca pts, lack of validation and standardization of specimens cross labs
9:44am May 1st 2018 via Twitter Web Client
Saddar: #ampeu2018 CRISPR and role in gene editing, they use it to create reference materials. Accuref is a div of Applied Stem Cell
Sonika Saddar (Accuref CA US) #ampeurope2018 Development and Characterization of Multigene Molecular Cytopathology… https://t.co/7T0Q7SceNV
9:42am May 1st 2018 via Twitter Web Client
Q: 5% and 1% in plasma is high; did the variant exist? Van Casteren: #ampeu2018 Some labs wanted to retest, but not… https://t.co/mf2Gqmsnu6
9:41am May 1st 2018 via Twitter Web Client
Q: Other metrics to compare labs? Yield, coverage? Van Casteren: #ampeu2018 They do collect it but haven't analyzed it, will expand it.
9:39am May 1st 2018 via Twitter Web Client
Q: Commentary on tech used? Van Casteren: #ampeu2018 asked each lab to treat as pt samples. Everything they current… https://t.co/K5UjzhQMtS
9:38am May 1st 2018 via Twitter Web Client
Van Casteren: #ampeu2018 Conclude - need further improvement needed for optimal pt treatment. Much higher FN rates relative to tissue.
Van Casteren: #ampeu2018 For different mut EGFR T790M by error type; main error is FNs in plasma.
9:35am May 1st 2018 via Twitter Web Client
Van Casteren: #ampeu2018 Looked at 5%, 1% VAF w/1 sample across 21; also tissue. 70% / 40% correct. 'Generally worse in plasma smpls'
9:33am May 1st 2018 via Twitter Web Client
Kaat Van Casteren (Leuven, Belgium) #ampeurope2018 External Quality Assessment Schemes for Gene Mutation Testing in… https://t.co/kYEpJAn3Q8
9:32am May 1st 2018 via Twitter Web Client
Hinerfeld: #ampeurope2018 Shows reprod data. 44-plex on FFPE lymphoid tissue, spatial structure and function.
8:55am May 1st 2018 via Twitter Web Client
Hinerfeld: #ampeurope2018 Internal to the DSP scope, a digital micromirror device (DMD) to select specific location… https://t.co/lLohU17QWf
8:51am May 1st 2018 via Twitter Web Client
Hinerfeld: #ampeurope2018 Able to do single-molecule counting; 40+ validated antibodies.
8:49am May 1st 2018 via Twitter Web Client
Hinerfeld: #ampeurope2018 and then the tags can be collected. They have a mock-up of the instrument in their exhibit.
Hinerfeld: #ampeurope2018 Protein reaents, RNA probes: a 4-channel microscope, stain and imaged. Can determine exac… https://t.co/gMVJGBHMfq
8:48am May 1st 2018 via Twitter Web Client
Hinerfeld: #ampeurope2018 Limited samples for IHC; Digital Spatial Profiling was then developed. More: https://t.co/tf5nE7XsjJ
8:46am May 1st 2018 via Twitter Web Client
Hinerfeld: #ampeurope2018 https://t.co/aUGnUQX0Xi '17 ref where it's not only about exp, but co-colocation of CD8, PD-L1
Doug Hinerfeld: #ampeurope2018 Powering precision oncology research: High plex multi-analyte profiling on FFPE w/Spatial Resolution
8:44am May 1st 2018 via Twitter Web Client
Q: Could it be possible to det the partner? van Kempen: #ampeurope2018 May be sufficient to know the driver for Rx
Q: 100 ng total RNA? What if less? van Kempen: #ampeurope2018 Counts may be too low, however felt it was feasible nonetheless.
8:43am May 1st 2018 via Twitter Web Client
Q: How does it work with a number of sample types?? van Kempen: #ampeurope2018 Manuscript under review; IHC concord 96% FISH 64%.
8:42am May 1st 2018 via Twitter Web Client
van Kempen: #ampeurope2018 Able to get down to 10% tumor cell % w/100ng total RNA input for ALK/RET/ROS1. 20% NRG1, 5%METx14
8:34am May 1st 2018 via Twitter Web Client
van Kempen: #ampeurope2018 100 ng total RNA from cell lines, get 20% LOD. Onto MET ex 14 skip, probe design, data.
8:33am May 1st 2018 via Twitter Web Client
van Kempen: #ampeurope2018 Due to high ROS1 endogenous expression, ROS1 meas noisy. NRG1 fusion design reviewed. Us… https://t.co/gqd1fwA1UW
8:29am May 1st 2018 via Twitter Web Client
van Kempen: #ampeurope2018 Example: EML4_6:ALK_20 fusion. Shows data from 65 pt samples, ROS1 signals via this method noisy
8:26am May 1st 2018 via Twitter Web Client
van Kempen: #ampeurope2018 NanoString Sprint: very easy, automated to use. Analysis - lots of numbers, use Excel to setup over-exp 3' end
8:24am May 1st 2018 via Twitter Web Client
van Kempen: #ampeurope2018 Shows even bigger list: "driven by expression, not mutation" (Bernards, earlier today) "Must have said it 20x"
8:20am May 1st 2018 via Twitter Web Client
van Kempen: #ampeurope2018 An imbalance probe design: 1:1 of WT; ratio >>1 of kinase to 5' partner.
8:19am May 1st 2018 via Twitter Web Client
van Kempen: #ampeurope2018 Designed panel for ALK/ROS1/RET/NTR1/NRG1/METex14skipping. Listed all the exons - many for each driver
8:18am May 1st 2018 via Twitter Web Client
van Kempen: #ampeurope2018 FISH for more DNA fusions means more slides. Diagram of NanoString approach '18 JMD https://t.co/rTtqzsi5H1
8:17am May 1st 2018 via Twitter Web Client
