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Illumina Cell-Free DNA Prep with Enrichment

A fast, flexible, and scalable cfDNA library preparation kit for highly sensitive mutation detection

~8.5–9.5 hr

Assay time

~2.5–3 hr

Hands-on time

10–30 ng of cfDNA

Input quantity

See full details in the specifications table

Overview

Illumina Cell-Free DNA Prep with Enrichment is a highly sensitive library preparation solution to detect low-abundance mutations from cfDNA samples. The workflow includes unique molecular identifiers (UMIs) for error correction and reduction of false positives, enabling accurate and sensitive detection of low-frequency mutations. Illumina Cell-Free DNA Prep with Enrichment has enhanced product attributes that enable greater efficiency for the labs, such as extended shelf life to six months and advanced change notification.

Fast, flexible format

Illumina Cell-Free DNA Prep with Enrichment is compatible with user-supplied enrichment oligo nucleotides from Illumina or third-party vendors, including single-stranded DNA (ssDNA) from Integrated DNA Technologies (IDT) and double-stranded DNA (dsDNA) from Twist Bioscience, for enhanced content portability. The kit accommodates 55-2000Kb (ssDNA) and 70-2000Kb(dsDNA) panel content, enabling flexible study design. Sequencing-ready libraries are prepared in ~8.5–9.5 hours, with only ~2.5-3 hours of hands-on time, enabling researchers to go from extracted cfDNA to sequencing in a single day. Recommended read length for NextSeq 550 is 2 × 149 bp. For NextSeq 2000, NovaSeq 6000, and NovaSeq X, the recommended read length is 2 × 151 bp.

High-performance library prep

Illumina Cell-Free DNA Prep with Enrichment detects low-frequency variants, including SNVs, CNVs, Indels as variants, and gene fusions. The kit identifies small variants with allele frequencies as low as 0.2% from only 20 ng cfDNA with ≥90% sensitivity.

Streamlined sample-to-insights workflow

This kit is part of an integrated workflow with option for no manual touchpoints from sequencer to insights that (1) starts with cfDNA, followed by (2) sequencing on Illumina mid- and high-throughput systems, (3) highly accurate variant calling using the DRAGEN for ILMN cfDNA Prep with Enrichment App, and (4) option for variant interpretation through automated draft research report generation using Illumina Connected Insights.

Flexible content options

Source your custom enrichment panel from Illumina, designed for your target gene list of interest. Design your custom panel easily with DesignStudio, a free online tool.

Learn more about cancer research applications

Specifications

Analytical sensitivity Analytical sensitivity at 20ng: ≥ 90% (SNVs ≥0.2% and indels ≥0.5%)
Analytical specificity Analytical specificity at 20ng: ≥ 99.98% (SNVs ≥0.2% and indels ≥0.5%)
Assay time ~8.5–9.5 hr
Hands-on time ~2.5–3 hr
Input quantity 10–30 ng of cfDNA
Instruments NextSeq 550 System, NextSeq 2000 System, NextSeq 1000 System, NextSeq 550Dx in Research Mode, NovaSeq X System, NovaSeq 6000 System
Method Custom sequencing, Targeted DNA sequencing, Target enrichment
Nucleic acid type DNA
Specialized sample types Cell-free DNA
Species category Human
Technology Sequencing
Variant class Gene fusions, Somatic variants, Single nucleotide variants (SNVs), Insertions-deletions (indels), Copy number variants (CNVs)

Required products

Index plates listed above are required to conplete the workflow. There are specific SKUs for manual and automation workflow. Illumina Custom Enrichment Panel v2 is a 120 bp, double-stranded probe panel designed using DesignStudio and includes enrichment oligos for hybridization capture.

/ Results

Applications

Example workflow

1
Prep
2
Sequence
3
Analyze

Variant calling

DRAGEN for ILMN cfDNA Prep with Enrichment

Insights and reporting

Illumina Connected Insights

Documentation

Product literature
Support documentation

Compare

Illumina Cell-Free DNA Prep with Enrichment Illumina DNA Prep with Enrichment TruSight Oncology 500 ctDNA v2
Analytical sensitivity Analytical sensitivity at 20ng: ≥ 90% (SNVs ≥0.2% and indels ≥0.5%) ≥ 95% (small variants, ≥ 0.5% VAF)
Analytical specificity Analytical specificity at 20ng: ≥ 99.98% (SNVs ≥0.2% and indels ≥0.5%) ≥ 99.995% (small variants, ≥ 0.5% VAF)
Assay time ~8.5–9.5 hr ~6.5 hr 3-4 days from purified nucleic acid to variant report
Hands-on time ~2.5–3 hr ~2 hr, ~2 hr ~1.5-2.5 hr for library prep and enrichment
Input quantity 10–30 ng of cfDNA 10-1000 ng high-quality genomic DNA or 50-1000 ng FFPE DNA. (For blood and saliva, see the reference guide). 20 ng cfDNA (4 ml of plasma)
Instruments NextSeq 550 System, NextSeq 2000 System, NextSeq 1000 System, NextSeq 550Dx in Research Mode, NovaSeq X System, NovaSeq 6000 System MiSeq System, iSeq 100 System, NextSeq 550 System, NextSeq 2000 System, MiSeqDx in Research Mode, MiniSeq System, NextSeq 550Dx in Research Mode, NovaSeq X System, NovaSeq 6000Dx in Research Mode, NovaSeq 6000 System, NovaSeq X Plus System NovaSeq 6000 System
Method Custom sequencing, Targeted DNA sequencing, Target enrichment Custom sequencing, Targeted DNA sequencing, Exome sequencing, Target enrichment Targeted DNA sequencing, Target enrichment
Nucleic acid type DNA DNA DNA
Specialized sample types Cell-free DNA Blood, Low-input samples, FFPE tissue, Saliva Circulating tumor DNA, Blood
Species category Human Other, Human Human
Technology Sequencing Sequencing Sequencing
Variant class Gene fusions, Somatic variants, Single nucleotide variants (SNVs), Insertions-deletions (indels), Copy number variants (CNVs) Single nucleotide polymorphisms (SNPs), Loss of heterozygosity (LOH), Somatic variants, Germline variants, Insertions-deletions (indels), Copy number variants (CNVs) Single nucleotide variants (SNVs), Insertions-deletions (indels), Copy number variants (CNVs)

*System compatibility: P4 and P3 flow cell tested

Resources

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Illumina® cfDNA Prep with Enrichment, Ligation (192 Samples, 4-plex), Cloud Analysis

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Includes ligation-based library prep and fast enrichment reagents for preparation of 192 libraries prepared by automation, or 240 libraries prepared manually, using 4-plex enrichment (pooling of 4 libraries in a single enrichment). Analyze data on Illumina Connected Analytics or BaseSpace Sequence Hub for free. Basic subscription to Cloud platform required. Data storage not included. UMI Index Adapters and Unique Dual Indexes are purchased separately and required for library preparation. Choose UMI Index Adapters and Unique Dual Indexes based on automation or manual workflow. Enrichment Panel Content not included.

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Panel (3)

Illumina Custom Enrichment Panel V2 (32 µl, 120 bp)

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Illumina Custom Enrichment Panel V2 enables 120 bp double-stranded custom panel design from 100 up to 1,000,000 probes using DesignStudio or Concierge Services and provides 32 µl of material (sufficient for 8 enrichment reactions when paired with Illumina DNA Prep with Enrichment). Library prep, enrichment, and index adapter reagents need to be ordered separately.

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Illumina Custom Enrichment Panel V2 (384 µl, 120 bp)

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Illumina Custom Enrichment Panel V2 enables 120 bp double-stranded custom panel design from 100 up to 1,000,000 probes using DesignStudio or Concierge Services and provides 384 µl of material (sufficient for 96 enrichment reactions when paired with Illumina DNA Prep with Enrichment). Library prep, enrichment, and index adapter reagents need to be ordered separately.

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Illumina Custom Enrichment Panel V2 (1536 µl, 120 bp)

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Accessory products (6)

IDT® for Illumina® UMI DNA/RNA UD Indexes Set A, Ligation (96 Indexes, 96 Samples)

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UMI DNA Index Anchors (Plate = 20027219, Box = 20032119) + Nextera Compatible Unique Dual Index A (Sales Kit = 20027213, Plate = 20025019, Box = 20026121)

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IDT® for Illumina® UMI DNA/RNA UD Indexes Set B, Ligation (96 Indexes, 96 Samples)

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IDT for Illumina UMI DNA Index Anchors Set B

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IDT for Illumina UMI DNA/DNA Index Anchors Set A for Automation

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Includes one box of 96 IDT for Illumina - UMI DNA/RNA UD Indexes Set A for Automation sufficient for labeling 96 samples and one box of 96 IDT for Illumina - UMI DNA Index Anchors for Automation.

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IDT for Illumina UMI DNA/DNA Index Anchors Set B for Automation

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Includes one box of 96 IDT for Illumina - UMI DNA/RNA UD Indexes Set B for Automation sufficient for labeling 96 samples and one box of 96 IDT for Illumina - UMI DNA Index Anchors for Automation.

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Illumina UMI DNA/RNA UD v3 indexes Set A, Ligation (96 indexes, 96 Samples)

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Includes one box of 96 Illumina DNA/RNA UD v3 Indexes Set A sufficient for labeling 96 samples and one box of 96 IDT for Illumina UMI DNA Index Anchors.

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Services (1)

Illumina Cell-Free DNA Prep with Enrichment Training - Customer site

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This 2-day introductory course is designed to familiarize new users with the Illumina cfDNA Prep with Enrichment workflow through detailed hands- on instruction. Participants will learn all essential steps in the workflow, including manual library preparation, enrichment, sequencing, and data analysis.

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