cfDNA Purification Kit (Magnetic Beads)

The cfDNA Purification Kit (Magnetic Beads) was developed for cell free DNA (cfDNA) enrichment by separating genomic DNA contamination from isolated cfDNA samples.

Many diagnostic technologies for detection of disease signals in cfDNA begin with isolation and purification of DNA from liquid biopsy that include urine, plasma, cerebrospinal fluid. The most widely explored biotechnology is assays used to detect cancer-derived plasma cfDNA. Silica-based magnetic bead cfDNA isolation kits can reliably extract total DNA from plasma, but typically yield a large variation in cfDNA that includes the presence of genomic DNA that often depends on tumor stage, tumor size, or healthy status individuals. Most of the commercial cfDNA isolation kits can’t specifically recover the cfDNA while leaving the high molecular weight genomic DNA behind. The presence of genomic DNA can lead to decreased sensitivity or inconsistent results in downstream applications such as next-generation sequencing (NGS), PCR, QPCR, and digital PCR etc.

Therefore, an additional purification step to enrich cfDNA before downstream methods helps to improve signal from fragments that originate from cancer cells. A proportion of cancer-derived cfDNA fragment signals are below 100 bp and are often not detectable except by qPCR or single-stranded DNA based library preparation for NGS (1, 2, 3). Furthermore only 1% of cancer-derived fragments are found above 400 bp (1, 2). Capture of size-selected fragments between 90-150 bp improved detection of cancer by 2-4 fold (4). Furthermore, TF-bound and protected cfDNA fragments are also being investigated for active cancer-specific signals down to 35-80 bp (5, 6).

This kit uses Dual Solid Phase Reversible Immobilization (SPRI) technology for cfDNA purification. Most Dual SPRI procedures do NOT recover fragments below 100 bp. The kit can be used for the enrichment of cfDNA isolated from liquid biopsies, plasma, serum, and urine. The kit separates cfDNA (50-500 bp) and genomic DNA, and recovers of 90% of the cfDNA without the high molecular weight genomic DNA with high efficiency. Fragments at 500 bp and above may also be retained. Both the 50-500 bp and >500 bp DNA fractions can be used for downstream applications such as single-stranded or double stranded NGS library prep, qPCR, ddPCR, and other methods.

Features

• Separation of cfDNA and genomic DNA; Recovery of both types of DNA
• Recovery of cfDNA (50-500 bp)
  • As short as 50 bp can be recovered
• Recovery of high molecular weight genomic DNA
• Removal of unwanted components and other impurities
• Automation friendly

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Examples of cfDNA purification. Both cfDNA and genomic DNA can be recovered separately.

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The range of recovered small DNA fragments is from 50 to 500 bp. The input DNA are mixtures of sheared small DNA fragments and intact genomic DNA. The ratios of sheared DNA fragments versus genomic DNA are indicated.

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Recovery rates of cfDNA and genomic DNA.

Many diagnostic technologies for detection of disease signals in cfDNA begin with isolation and purification of DNA from liquid biopsy that include urine, plasma, cerebrospinal fluid. The most widely explored biotechnology is assays used to detect cancer-derived plasma cfDNA. Silica-based magnetic bead cfDNA isolation kits can reliably extract total DNA from plasma, but typically yield a large variation in cfDNA that includes the presence of genomic DNA that often depends on tumor stage, tumor size, or healthy status individuals. Most of the commercial cfDNA isolation kits can’t specifically recover the cfDNA while leaving the high molecular weight genomic DNA behind. The presence of genomic DNA can lead to decreased sensitivity or inconsistent results in downstream applications such as next-generation sequencing (NGS), PCR, QPCR, and digital PCR etc.

Overview

• Detection kits for Candida albicans
• CE-IVD marked version available for in vitro diagnostic use
• Available in TaqMan format for analysis

Candida albicans is an opportunistic yeast and is the most common fungal pathogen found in the human body. C. albicans can be detected in the gastrointestinal tract, mouth, and vagina of approximately half of healthy adults. It is typically a commensal organism and makes up part of the natural human microflora; however it can overgrow and become pathogenic as a result of various conditions. For example, individuals who have taken recent courses of antibiotics, have a weakened immune system or have diabetes have an increased risk of developing a Candida albicans infection. When an overgrowth occurs, this can lead to common infections such as urinary tract infections, genital yeast infections, oral thrush, and mucocutaneous candidiasis. In the more severe cases, when Candida albicans enters the bloodstream or organs, it can lead to loss of sight, blood and bone infections, endocarditis, meningitis or inflammation of the intraabdominal lining.

Candida albicans TaqMan PCR Kit, 100 reactions

• Ready to use format, including Master Mix for the target and PCR control to monitor for PCR inhibition and validate the quality
• Specific Primer and Probe mix for the pathogen/virus/viroid of interest
• Primer and Probe mix
• Positive and negative control to confirm the integrity of the kit reagents

Candida albicans TaqMan PCR Probe/Primer Set and Controls, 100 reactions

• Specific Primer/Probe mix and Positive Control for the pathogen/virus/viroid of interest
• Nuclease-free water
• Can be used together with Norgen’s PCR Master Mix (#28007) or customer supplied master mix

Details

Supporting Data

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Storage Conditions and Product Stability
All kit components can be stored for 2 years after the date of production without showing any reduction in performance.

All kit components should be stored at -20°C upon arrival. Repeated thawing and freezing (> 2 x) of the Master Mix and Positive Control should be avoided, as this may affect the performance of the assay. If the reagents are to be used only intermittently, they should be frozen in aliquots.

Component	Cat. TM34050 (100 preps)	Cat. TM34010 (100 preps)
MDx TaqMan 2X PCR Master Mix	2 x 700 μL	–
Candida albicans Primer & Probe Mix	280 μL	280 μL
Candida albicans Positive Control	150 μL	150 μL
Nuclease-Free Water (Negative Control)	1.25 mL	1.25 mL
Product Insert	1	1

Introduction

Usages:
For differentiating enteric bacteria based on urease activity by adding 40% sterile urea solution.

Principle:
Peptone provides the carbon and nitrogen; maintain a balanced osmotic sodium chloride; potassium dihydrogen phosphate is buffers; decomposing bacteria urease urea medium, produce large amounts of ammonia, agar as medium coagulant.

Formulation (per liter):
Peptone 1g
Sodium chloride 5g
Glucose 1g
Ppotassium dihydrogen phosphate 2g
Phenol red 0.012g
Agar 12g
Final pH7.2 ± 0.2

How to use:
1.Suspend 21g in 1L of distilled or deionized water. Heat with frequent agitation and boil to completely dissolve the powder. Distribute into flasks. Autoclave at 121 for 15 minutes. cooling to 50-55 and adding 40% urea solution.
2.Diluted and treated samples.

Storage: Store in a dark, cool and dry place, tighten the cap immediately after use. Storage period of three years.

500g