Panel-Plus and CodeSet-Plus
Contents
1. Expression 100000 Human reference total RNA 100 pg was converted to cDNA and enriched via a Multiplexed Target Enrichment MTE with ee primers appropriate for downstream hybridization according er to the nCounter Single Cell Expression Assay Protocol for o Total RNA The enriched cDNA was hybridized with either the 100 gt nCounter Cancer Reference Kit alone Original CodeSet or ag with the nCounter Cancer Reference Kit and a 30 gene Panel 1 Plus Original CodeSet with Plus Assays were performed in triplicate for each condition tested and normalized to internal Cancer Reference Panel Only 10 100 1000 10000 10000 positive controls Average count values for each triplicate Cancer Reference Panel with 30 gene Plus Product measurement were determined and plotted for all nCounter Cancer Reference Kit probes in the original CodeSet alone y axis and Original CodeSet with Plus x axis Design and Ordering Information When a custom nCounter Panel Plus or CodeSet Plus is ordered probes are a designed for every gene sequence in the request Two separate vials for custom Description Number of Genes Number of Reactions Plus genes are shipped along with a unique Add in Library File ALF file that Panel Plus 6 11 genes 48 minimum includes the additional genes The Plus products will not include controls but 11 20 genes are tested prior to shipment with the nCounter standard CodeSet controls 21 30 genes i2. be sent within 24 hours 4 How do l analyze the combined data After scanning on the Digital Analyzer data may be imported into nSolver Analysis Software for downstream analysis Please see the nSolver Analysis Software Manual for additional information on the nSolver application 5 Can I purchase a Panel Plus for my miRNA miRGE or Plex2 experiment Not at this time Plus products are specially formulated to be universally compatible with all CNV and Gene Expression CodeSets NanoString Technologies Inc 530 Fairview Ave N LEARN MORE SALES CONTACT Suite 2000 To learn more about Panel Plus and CodeSet Plus United States us sales nanostring com Seattle WA 98109 USA products visit http www nanostring com products Europe europe sales nanostring com panel_plus or contact your local NanoString Technologies representative Toll free 1888 358 6266 Fax 206 378 6288 Other Regions info nanostring com www nanostring com info nanostring com 2014 NanoString Technologies Inc All rights reserved NanoString NanoString Technologies nCounter Molecules That Count and nSolver are registered trademarks or trademarks of NanoString Technologies Inc NanoString in the United States and or other countries All other trademarks and or service marks not owned by NanoString that appear in this document are the property of their respective owners The manufacture use and or sale of NanoString product s
3. may be subject to one or more patents or pending patent applications owned by NanoString or licensed to NanoString from Life Technologies Corporation and other third parties FOR RESEARCH USE ONLY Not for use in diagnostic procedures NOVEMBER 2014 v 20141104
4. d flexibility to tailor a comprehensive list of genes to your exact needs In addition combining a panel kit combined with Panel Plus will allow you to survey a comprehensive collection of genes for a lower cost per sample when compared to the same gene set being ordered as a custom CodeSet CodeSet Plus provides researchers the ability to update custom CodeSets that may be used to analyze large sample sets over many months Genes of interest that become known during ongoing research or from a new publication can now easily be added to a custom CodeSet This flexibility means that a custom CodsSet sufficient to analyze all the samples in a large study can be purchased with confidence Purchasing a larger custom CodeSets provides a more cost effective solution versus purchasing multiple smaller custom CodeSets Molecules That Count Translational Research Gene Expression miRNA Expression Epigenomics CNV Single Cell 1 FRODUCIT DAWA Sree nCounter Gene Expression and CNV Panel Plus and CodeSet Plus nCounter Panel Plus and CodeSet Plus Product Overview Plus CodeSets contain Reporter Reporter Plus and Capture Capture Plus Probes for up to 30 targets and can be combined with any custom CodeSet or pre designed Panel prior to overnight hybridization Using a Plus product with either a CodeSet or Panel is simple and requires few additional steps as outlined in the diagram below k NOTE After hybridization all subsequent steps are performed a
5. nCounter Gene Expression and CNV Panel Plus and CodeSet Plus PRODUCT IAW Slee If nN a N S t r nN S t E C HN OLr rO GIE S Panel Plus and CodeSet Plus Product Highlights e 30 unique Reporter Probes are universally compatible with all custom CodeSets and Panels e Compatible with all Gene Expression and CNV products including Single Cell applications Applications e Customize a Panel Add up to 30 of your favorite genes ora collection of specific controls e Update a large CodeSet Add new genes as you go to keep your CodeSet updated e Create specific control sets Create multiple Plus control sets for use with different sample or cell types e Finalize your gene list more quickly Add more genes later nCounter Panel Plus and CodeSet Plus Product Overview The nCounter Panel Plus and CodeSet Plus products from NanoString Technologies add a new level of flexibility to nCounter Gene Expression and CNV experiments Researchers can now customize any NanoString off the shelf panel kit or add genes to any custom CodeSet by utilizing up to 30 unique Reporter Probes exclusively formulated for use in Panel Plus and CodeSet Plus products Panel Plus enables any off the shelf panel kit to be customized with up to 30 additional genes of interest Customizing a panel kit with a Panel Plus product allows researchers to leverage the bioinformatics knowledge incorporated into every nCounter panel kit and provides unprecedente
6. ncluded in all custom CodeSets and panels Plus products are always used with either a panel or a custom CodeSet and do not require the purchase of poco Flus a ee 48 minimum additional Master Kit reagents U genes 21 30 genes Plus products can be ordered from three separate gene categories and are offered with purchases of Panels or Custom CodeSets for 48 reactions or more Molecules That Count 3 PRODUCT IAW Slee nCounter Gene Expression and CNV Panel Plus and CodeSet Plus Frequently Asked Questions 1 Can I use multiple Plus products with a Panel or Custom CodeSet No Multiple Plus products cannot be mixed or combined in a reaction Only one Plus product may be used in conjunction with a set of specific reactions It is possible to use different Plus products on separate occasions with different Panels or CodeSets 2 Can luse the Plus product as would a custom CodeSet No Plus products are not the same as custom CodeSets Plus products have been formulated to be mixed with Panel or Custom CodeSet Reporter and Capture Probe Mixes Plus products also do not include the necessary controls for proper normalization 3 How dol include the new genes in my data analysis To include the new genes please contact NanoString Bioinformatics at bioinformatics nanostring com Include in your message the name of the Plus product and original CodeSet that will be combined A new Reporter Library File RLF containing both will
7. pL to Capture ProbeSet 4 Complete Hybridization Reactions To each tube add 6 uL Capture ProbeSet and Plus Reagent Mix 4 a 4 lt q 1 Cap tube mix by inverting strip 2 Briefly spin down 3 Incubate at 65 C 33 40 pL total volume after Capture Probe nanostring 2 T E C HN OF LOGIES NanoString Technologies PRODUCT DATA SHEET Performance of Panel Plus Gene Sets with Pre designed Panels Standard Gene Expression and Single Cell Analysis Two experiments were performed by adding Panel Plus genes to the Human Cancer Reference Kit Data in each figure below demonstrate that addition of CodeSet Plus probes does not impact performance of original nCounter panel probes or single cell panel probes Gene Expression 2 100000 Human reference total RNA 100 ng was hybridized with either O caer the nCounter Cancer Reference Kit alone Original CodeSet or z aa with the nCounter Cancer Reference Kit and a 30 gene Panel a 1000 Plus Original CodeSet with Plus Assays were performed in at triplicate for each condition tested and normalized to internal S 100 v positive controls Average count values for each triplicate g ic measurement were determined and plotted for all nCounter D Cancer Reference Kit probes in the Original CodeSet alone 1 y axis and Original CodeSet with Plus x axis O 10 100 1000 10000 10000 Cancer Reference Panel with 30 gene Plus Product Single Cell Gene
8. s described in the nCounter Prep Station and Digital Analyzer Manuals For specific recommendations on sample input see the nCounter RNA or DNA User Manual For a detailed protocol see the nCounter Panel Plus and CodeSet Plus Protocol 1 Create Reporter and Plus Reagent Master Mix r 4 Add 26 uL to Reporter CodeSet 1 r Add 130 uL to Reporter CodeSet I I I I l l I l i Yy y Reporter Plus Reagent Hybridization Buffer Reporter CodeSet MasterMix Thaw 12 reactions on ice Invert to mix Invert to mix and spin down Thaw 12 reactions on ice Invert to mix and and spin down Invert to mix and spin down spin down Sanma E Add 26 uL to Reporter CodeSet _f S5888o Add 130 pL to Reporter CodeSet 2 Set Up Hybridization Reaction To each tube add 27 uL is the standard Gene Expression and CNV hybridization set up volume Up to 37 uL can be used for Single Cell assays that include MTE 22 uL Master Mix volumes up to 15 uL per sample I I I I RNAse free Water I if needed I I I I Minimum 27 pL total volume per tube 3 Create Capture ProbeSet and Plus Reagent Mix r Add 14 uL to Capture ProbeSet l 1 y Capture Plus Reagent Capture ProbeSet Capture ProbeSet and Plus Reagent Mix Thaw 12 reactions on ice Invert to mix and Thaw 12 reactions on ice Invert to mix and spin down spin down MS I Add 14
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