To accommodate the need for high quality enzymes for isothermal amplification ArcticZymes developed the IsoPol® series of polymerases. For isothermal amplification at lower temperatures (37°C and below) we offer two enzymes, IsoPol® DNA Polymerase and IsoPol® SD+. Both enzymes exhibit excellent processivity and high strand displacement activity, having 5’-3’ polymerase activity while lacking both 3’-5’and 5’-3’ exonuclease activity.
Detail
OverView
To accommodate the need for high quality enzymes for isothermal amplification ArcticZymes developed the IsoPol® series of polymerases. For isothermal amplification at lower temperatures (37°C and below) we offer two enzymes, IsoPol® DNA Polymerase and IsoPol® SD+. Both enzymes exhibit excellent processivity and high strand displacement activity, having 5’-3’ polymerase activity while lacking both 3’-5’and 5’-3’ exonuclease activity.
IsoPol® SD+ is an engineered version of IsoPol® DNA Polymerase with even stronger strand displacement and higher salt tolerance.
Figures
Properties
Quality Control
ArcticZymes is dedicated to the quality of our products. IsoPol® polymerases are is manufactured at our ISO 13485 certified facility in Norway.
Other Products
Cat.# 20104S, 20104L: Size range 250-350 bp (ideal for NGS library size selection)
Product Info
Document
Product Info
The series of DNA Size Selection Kits (Magnetic Beads) were developed for DNA size selection using magnetic beads. A total of 11 kits are available, with different selection ranges spanning from 50 bp to over 10 kb. The kits provide a simple and quick approach for the enrichment of a specific range of DNA fragments. The kit workflow allows double-sided or single-sided size selection for specific size cutoffs.
Gel images of different ranges of size selection. Sheared human genomic DNA was used as input.
.
DNA size selection is a selective capture of DNA fragments of a specific range of size for next-generation sequencing (NGS) library preparations, PCR, ChIP assay, DNA ligations, endonuclease digestions, adapter removal, and other genomics and molecular biology applications. DNA size selection is preferred after NGS library prep in most of the cases. The NGS library preparation is related to the quality of the sequencing data. Precise NGS library size selection can increase sequencing efficiency, improve data quality, and reduce costs.
There are two types of sequencing technologies: short-read sequencing and long-read sequencing. Short-read sequencing uses DNA libraries that contain small insert DNA fragments of similar sizes, usually several hundred base pairs. The sequencing efficiency can be improved if the DNA size selection is in the right range. Cat.# 20104S and 20104L are the best kits for NGS library size selection of illumina paired-end 100 (PE100) sequencing with 100-200 bp library inserts; Cat.# 20105S and 20105L are the best kits for NGS library size selection of illumina paired-end 150 (PE150) sequencing with 150-300 bp library inserts; and Cat.# 20106S and 20106L are the best kits for NGS library size selection of illumina paired-end 300 (PE300) sequencing with 300-600 bp library inserts.
Long-read sequencing uses a large DNA fragment as input and makes very long reads. Usually, library size selection is preferred to remove smaller fragments. Cat.# 20110S and 20110L are the best kits for long-read sequencing size selection with DNA sizes >5 kb, and Cat.# 20111S and 20111L are the best kits for long-read sequencing size selection with DNA sizes >10 kb.
The magnetic beads technology uses paramagnetic particles, also known as SPRI (Solid Phase Reversible Immobilization) beads, to bind DNA reversibly and selectively. DNA fragments can be size-selected and purified by changing the properties of the magnetic beads or SPRI beads. The magnetic beads can easily separate the beads-binding DNA from the contaminants and unwanted components in the samples. The samples after DNA size selection are free of contaminants such as buffer components, enzymes, proteins, salts, dNTPs, primers, and adapters. Our proprietary magnetic beads reagents improve yield, selectivity, and reproducibility.
Specific DNA fragments at a certain length range can be purified simply using magnetic separation with different beads components, avoiding tedious and time-consuming gel extraction and column-based purification. The magnetic beads method is popular for common DNA size selection, including library size selection. The first beads-binding step, referred to as the right-side clean-up, removes large DNA fragments. The large DNA fragments are bound to the beads and are discarded. The desired DNA fragments in the supernatant are transferred to a new well, and new beads are added to the supernatant for the second beads-binding, referred to as the left-side clean-up. The double-size selected DNA fragments are eluted after ethanol rinsing.
DNA size selection with dual clean-ups.
.
A single clean-up is needed for DNA size selection with large fragments. In this case, only the large DNA fragments are bound to the beads. The selected larger DNA fragments are eluted after ethanol rinsing.
DNA size selection with single clean-up for >5 kb and >10 kb DNA.
.
Features of DNA size selection and library size selection
High specificity and high recovery of size selection
11 selection ranges are available, including 5 ranges for NGS library size selection
50-100 bp
100-200 bp
200-500 bp
250-350 bp: ideal for illumina PE100 sequencing
300-450 bp: ideal for illumina PE150 sequencing
450-750 bp: ideal for illumina PE300 sequencing
500-1000 bp
1-3 kb
1-5 kb
>5 kb: ideal for long-read sequencing
>10 kb: ideal for long-read sequencing
Fast and simple
20-min protocol
No gel purification required
No columns required
No centrifugation required
Efficient removal of contaminants and unwanted components
CE-IVDR marked in accordance with the European Commission Regulation (EU) No. 2017/746.
Ideal for use in in vitro diagnostic workflows
Reliable and cost-effective
Non-invasive, user-friendly sample collection
Sample collection and preservation in one convenient kit
Samples are non-infectious and can be handled and shipped safely
Preserved DNA is stable for 2 years at ambient temperature
High quality DNA is suitable for sensitive downstream applications including PCR, qPCR, sequencing, SNP analysis, microarrays, RFLP and Southern Blot Analysis
Shipping accessories can be purchased separately
Norgen’s Saliva DNA Collection and Preservation Devices are designed for 1) simple and non-invasive saliva collection and 2) preservation of DNA in saliva samples at ambient temperature. Each of the 50 Saliva DNA Collection and Preservation Devices consists of 3 components: (1) Saliva Collection Funnel and Collection Tube, (2) Collection Tube Cap, and (3) Norgen’s Saliva DNA Preservative contained within a sealed squeezable ampoule. Saliva samples are collected by spitting inside the Collection Funnel which has been assembled with the Collection Tube. After collecting the required volume of saliva the Collection Funnel is removed and the contents of the Preservative Ampoule are then added and mixed with the collected saliva. The Saliva Collection Tube is subsequently sent to the laboratory for DNA isolation and analysis. DNA can be isolated from the preserved saliva samples using Norgen’s Saliva DNA Isolation Kit (Cat# Dx45400). Each of Norgen’s Collection Tubes is labeled with a unique serial number that can be used for secure and anonymous tracking of the sample. The saliva DNA in preserved samples is stable for more than 2 years at room temperature. This kit is ideal for collecting and preserving DNA samples for in vitro diagnostic use for medical purposes.
Saliva DNA Preservative
Norgen’s Saliva DNA Preservative is an aqueous storage buffer designed for rapid cellular lysis and subsequent preservation of DNA from fresh specimens. The buffer prevents the growth of Gram-negative and Gram-positive bacteria and fungi, and also inactivates viruses allowing the resulting non-infectious samples to be handled and shipped safely. In addition, the buffer eliminates the need to immediately process or freeze samples and allows the samples to be shipped to centralized testing facilities at ambient temperature. The components of the buffer allow samples to be stored for more than 2 years without any detectable DNA degradation.
DNA Isolation from Preservative
Prior to saliva DNA isolation, vortex the Collection Tube containing preserved saliva for 10 seconds and incubate at 55°C for one hour. Saliva DNA can now be isolated from the preserved saliva samples using Norgen’s spin-column based Saliva DNA Isolation Kit Dx (Cat# Dx45400) or other commercially available methods that have been validated for use with Norgen’s Saliva DNA Collection and Preservation Devices.
NOTE: This product is not available for sale in the United States.
The Collection Device should be kept tightly sealed and stored at room temperature for up to 3 years without any reduction in kit performance (The collection due date is written on the device label).
Once collected, saliva is stable for more than 2 years when kept tightly sealed and stored at room temperature.
The Collection Tube, the Collection Funnel and the Device Container of each Individual Saliva DNA Collection and Preservation Device are recyclable.
Kit Components
Cat. 49000 (50 Devices)
Individual Saliva DNA Collection and Preservation Devices
50
Donor Procedure Flowchart
1
Product Insert
1
Individual Saliva DNA Collection and Preservation Device Contents
Apoptosis is an essentially normal physiological process that removes now redundant, cells, particularly during embryonic development and early growth. In adult animals the process removes cells that are irreparable. The apoptotic process is also involved in many major diseases such as cancer, where transformed tumour cells have their apoptotic process disabled, permitting cell cycling to continue unchecked. In contrast some forms of senile dementia may result from excessive apoptotic induction of neural cells.
The apoptotic process in mammalian cells is a rapid event (2‐4 hours). Within this short time span an apparently viable cell can be quietly dismantled, to disappear leaving no visible trace of its former existence.
How is apoptosis detected or measured?
An apoptosis cascade of activators, effectors and regulators has been identified. This in turn led to a range of apoptosis assays being devised to detect and monitor these events. Some laboratories will employ two distinct assays, one selected to detect early (initiation) apoptotic events, while a second assay will target a later (execution) event. Apoptosis assays, based on methodology, can be classified into four major inter‐linked groups:
[1] DNA fragmentation (electrophoresis and nick end labelling, TUNEL).
[2] Apoptotic proteases (fluorescently labelled antibodies to the caspases).
[3] Flow cytometric analysis (FACS, incorporating other group assays).
Biocolor’s APOPercentage assay is based on the latter. Further information can be found under the ‘Mode of Action’ Tab.
How does APOPercentage detect apoptosis?
The mammalian cell membrane has been described as a semi‐fluid mosaic structure, composed of phospholipids with a diverse group of inserted proteins and some cholesterol. The phospholipids are the major components of the membrane and are arranged in the form of a ‘bi‐layer’; which is asymmetric in composition, structure, and function.
To ensure normal transmembrane functions the phospholipids must be maintained in an asymmetric composition. The process is regulated by ‘flippases’, which catalyse the active transport of aminophospholipids from the outer to inner monolayer. However, in cells undergoing apoptosis, flippase is overwhelmed by the action of another enzyme, termed ‘floppase’ or ‘scramblase’. The net effect is a scrambling of the phospholipid distribution between the inner and outer monolayers.
Cell membrane changes during apoptosis
The APOPercentage assay utilises an intense, pink-coloured dye reagent which is taken up during in-vitro culture by apoptosis-committed cells. This uptake occurs at the stage of Phosphatidylserine transmembrane movement, as produced by the flipflop mechanism. Dye uptake continues until blebbing occurs. No further dye can then enter the now defunct cell and the dye that has accumulated within the cell is not released (unlike necrotic cells which release dye).
Since the dye reagent is excluded or not retained by healthy or necrotic cells it therefore acts as a specific label for apoptotic cells.
How are APOPercentage-labelled cells quantified?
Labelled apoptosis cells may then by conveniently analysed by the following methods:
Direct Analysis The intense pink colour of the labelled cells can be visually assessed using brightfield microscopy. Apoptosis in substrate-adherent cell populations is therefore readily quantified using image analysis techniques. This technique is the most sensitive with the ability of detecting one single apoptotic cell per well.
Colorimetry protocol Dye that accumulates within apoptotic cells is released into solution via addition of Dye Release Reagent. The concentration of this intracellular dye is then measured at 550nm using a microplate colorimeter/spectrophotometer.
NB: The APOPercentage assay kit does NOT require the use of a Flow Cytometer.
Limit of Detection
A single cell (via image analysis method)
Detection Method
Colorimetric (550nm) (Endpoint) or Image Analysis based
Measurements per kit
Sufficient for 4×24 well plates or 6×96 well plates
Suitable Samples
Adherent mammalian cells (in-vitro)
APOPercentage kit contents:
1. APOPercentage Dye (1x5ml)
2. Dye Release Reagent (1x150ml)
3. Phosphate Buffered Saline (PBS) (1x120ml)
4. 24-well starter plate.
5. Assay kit manual.
The Colorimetric Protocol requires a Microplate Colorimeter / Spectrophotometer.
Additional 96-well plates will be required for use when reading dye absorbance values.
The Direct Detection Protocol Requires an inverted stage microscope with an attached digital camera.
NB: Additional reagents (typically culture medium and suitable apoptosis treatments) may be required for sample preparation prior to assay. Consult manual or contact us for further details.
Document
The APOPercentage™ Apoptosis kit is a dye-based, colorimetric assay for detection and measurement of apoptosis (programmed cell death) during in-vitro cell culture.
