Introduction

Free-circulating nucleic acids, such as tumor-specific extracellular DNA fragments and mRNAs in the blood or fetal nucleic acids in maternal blood, are present in serum or plasma usually as short fragments, <1000bp (DNA). HiPure Circulating DNA Midi Kit enables efficient purification of these circulating nucleic acids from human plasma, serum, or urine. The extracted products can be used for clinical in vitro detection.

Details

Specifications

Features	Specifications
Main Functions	Isolation circulating DNA from 1-5ml plasma, serum, body fluids using vacuum protocol
Applications	qPCR, liquid or solid chip analysis, hybridization and SNP detection, etc.
Purification method	Mini spin column
Purification technology	Silica technology
Process method	Manual (vacuum)
Sample type	Serum, plasma and other cell-free fluid samples
Sample amount	1-5ml
Elution volume	≥50μl
Time per run	≤60 minutes
Liquid carrying volume per column	4ml
Binding yield of column	1mg

Principle

This product is based on silica Column purification. The sample is lysed and digested with lysate and protease, DNA is released into the lysate. Transfer to an adsorption column. Nucleic acid is adsorbed on the membrane, while protein is not adsorbed and is removed with filtration. After washing proteins and other impurities, Nucleic acid was finally eluted with low-salt buffer.

Advantages

• High yield – most optimal process, free DNA (>50bp) can be obtained to the maximum extent
• High concentration – low elution volume, ensuring high nucleic acid concentration
• High purity – low alcohol binding method, completely removing inhibitor and protein pollution
• High recovery – DNA can be recovered at thelevel of PG by silica gel column purification

Kit Contents

Contents	IVD3182
Purification Times	50
Buffer ACL	250 ml
Buffer ACB*	300 ml
Buffer DCW1*	22 ml
Buffer DCW2*	10 ml
Proteinase K	540 mg
Protease Dissolve Buffer	30 ml
Carrier RNA	110 μg
Nuclease Free Water	20 ml
HiPure CFDNA Mini Columns	50
2 ml Collection Tubes	100
Extender Tube	50
Vac-Connector	50

Storage and Stability

Proteinase K, Carrier RNA should be stored at 2-8°C upon arrival. However, short-term storage (up to 12 weeks) at room temperature (15-25°C) does not affect their performance. The remaining kit components can be stored dry at room temperature (15-25°C) and are stable for at least 18 months under these conditions. The entire kit can be stored at 2-8°C, but in this case buffers should be redissolved before use. Make sure that all buffers are at room temperature when used.

Experiment Data

Free-circulating nucleic acids, such as tumor-specific extracellular DNA fragments and mRNAs in the blood or fetal nucleic acids in maternal blood, are present in serum or plasma usually as short fragments,

Introduction

The Kit provides fast purification of high-quality RNA from paxigene Blood RNA Tube using silica-membrane spin columns. RNA purified using the HiPure System is ready for applications such as RT-PCR, Northern blotting, poly A+ RNA (mRNA) purification, nuclease protection, and in vitro translation.

Details

Specifications

Features	Specifications
Main Functions	Isolation total RNA from Paxgene RNA Tubes
Applications	RT-PCR, qPCR, Northern hybridization, NGS, nucleic acid protection, in vitro translation
Purification method	Mini spin column
Purification technology	Silica technology, DNA filtration technology
Process method	Manual (centrifugation or vacuum)
Sample type	Blood in the preservation tube
Sample amount	2.5ml
Elution volume	≥30μl
Liquid carrying volume per column	800µl
Binding yield of column	100μg

Principle

The Kit is for the purification of total RNA from 2.5 ml human whole blood collected in a PAXgene Blood RNA Tube. Purification begins with a centrifugation step to pellet nucleic acids in the PAXgene Blood RNA Tube. The pellet is washed and resuspended, and incubated in optimized buffers together with proteinase K. DNA wash The lysate is passed through a DNA Mini column. Ethanolis added to adjust binding conditions, and the lysate is applied to a column.RNA is selectively bound to the silica membrane as contaminants pass through. Remaining contaminants are removed in several efficient wash steps. Between the first and second wash steps, the membrane is treated with DNase I to remove trace amounts of bound DNA. After the wash steps, RNA is eluted in elution buffer and heat-denatured.

Advantages

• High quality – high purity total RNA can be directly used in various sensitive downstream applications
• Fast – isolation of several samples can be completed in 40 minutes by using column purification method
• Safety – no phenol chloroform extraction required
• Sensitive – direct lysis of blood, plasma and other samples without separation of leukocytes

Kit Contents

Contents	R416802	R416803
Purification Times	10 Preps	50 Preps
HiPure RNA Mini Columns I	10	50
gDNA Filter Mini Columns	10	50
2ml Collection Tubes	30	150
RNase Free Water	60 ml	250 ml
Buffer MBR1	10 ml	30 ml
Buffer MBR2	5 ml	15 ml
Buffer RW1	15 ml	60 ml
Buffer RW2*	6 ml	20 ml
Proteinase K	12 mg	50 mg
Protease Dissolve Buffer	1.8 ml	5 ml
DNase I	120 µl	600 µl
DNase Buffer	6 ml	30 ml

Storage and Stability

Proteinase K should be stored at 2–8°C upon arrival. DNase I should be stored at -20°C. However, short-term storage (DNase I up to 1 weeks, Proteinase K up to 8 weeks) at room temperature (15–25°C) does not affect their performance. The remaining kit components can be stored at room temperature (15–25°C) and are stable for at least 18 months under these conditions.

Experiment Data

The Kit provides fast purification of high-quality RNA from paxigene Blood RNA Tube using silica-membrane spin columns. RNA purified using the HiPure System is ready for applications such as RT-PCR, Northern blotting, poly A+ RNA (mRNA) purification, nuclease protection, and in vitro translation.

Overview

• Extract total RNA (including microRNA) from FFPE samples
• No phenol extraction step
• Includes DNase for optional on-column DNA removal
• Isolated RNA is of the highest quality and integrity
• Isolate a diversity of RNA species
• Purified RNA is suitable for a variety of downstream applications, including Small RNA Sequencing. Find out more information on Norgen’s NGS services
• Purification is based on spin column chromatography that uses Norgen’s proprietary resin separation matrix

Norgen’s FFPE RNA Purification Kits provide a rapid method for the isolation and purification of total RNA (including microRNA) from formalin-fixed paraffin-embedded (FFPE) tissue samples in as little as 1 hour. Using formalin to fix tissues leads to crosslinking of the RNA and proteins, and the process of embedding the tissue samples can also lead to fragmentation of the RNA over time. Norgen’s FFPE RNA Purification Kits provide conditions that allow for the partial reversing of the formalin modifications, resulting in a high quality and yield of RNA. These kits are able to purify all sizes of RNA, from large mRNA and ribosomal RNA down to microRNA (miRNA) and small interfering RNA (siRNA), depending on the age of the FFPE tissue as fragmentation of the RNA is known to occur over time. The RNA is preferentially purified from other cellular components without the use of phenol or chloroform.

FFPE RNA Purification Kit (Spin Column)

Maximum loading volume of 650 μL per column, and a maximum binding capacity of 50 μg per column.

FFPE RNA Purification 96-Well Kit (High Throughput)

Purification is based on 96-well column chromatography using Norgen’s proprietary resin as the separation matrix. Purification can be performed using either a vacuum manifold or centrifugation. Maximum loading volume of 400 μL per well, and a maximum binding capacity of 50 μg per well.

Details

Supporting Data

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Kit Specifications
Maximum Binding Capacity Per Well	50 μg
Maximum Loading Volume Per Well	400 μL
Size of RNA Purified	All sizes, including small RNA (< 200 nt)
Maximum Amount of Starting Material:	5 slices of < 20 µm thick paraffin  25 mg of unsectioned block

Storage Conditions and Product Stability
All solutions should be kept tightly sealed and stored at room temperature. The DNAse I should be stored at -20°C upon arrival. The Proteinase K should be stored at -20°C upon arrival and after reconstitution. This kit is stable for 1 year after the date of shipment.

Component	Cat. 25300 (50 preps)	Cat. 25400 (2 x 96 preps)
Digestion Buffer A	25 mL	2 x 25 mL
Buffer RL	30 mL	2 x 30 mL
Enzyme Incubation Buffer	6 mL	2 x 6 mL
Wash Solution A	38 mL	2 x 38 mL
Elution Solution A	6 mL	2 x 20 mL
Proteinase K	12 mg	2 x 20 mg
DNase I	1 vial	2 x 500μL
Micro Spin Columns	50	–
96-Well Incubation Plate	–	2
96-Well Plate	–	2
Adhesive Tape	–	8
Collection Tubes	50	–
96-Well Collection Plate	–	2
Elution Tubes (1.7 mL)	50	–
96-Well Elution Plate	–	2
Product Insert	1	1