Description

Escherichia coli is one of many species of bacteria living in the lower intestines of mammals, known as gut flora. When located in the large intestine, it assists with waste processing, vitamin K production, and food absorption. Discovered in 1885 by Theodor Escherich, a German pediatrician and bacteriologist, E. coli are abundant: the number of individual E. coli bacteria in the faeces that a human defecates in one day averages between 100 billion and 10 trillion. However, the bacteria are not confined to the environment, and specimens have also been located, for example, on the edge of hot springs. The E. coli strain O157:H7 is one of hundreds of strains of the bacterium that causes illness in humans.

E. coli are unable to sporulate. Thus, treatments which kill all active bacteria, such as pasteurization or simple boiling, are effective for their eradication, without requiring the more rigorous sterilization which also deactivates spores. As a result of their adaptation to mammalian intestines, E. coli grow best in vivo or at the higher temperatures characteristic of such an environment, rather than the cooler temperatures found in soil and other environments.

The enteric E. coli (EC) are divided on the basis of virulence properties into enterotoxigenic (ETEC – causative agent of diarrhea in humans, pigs, sheep, goats, cattle, dogs, and horses), enteropathogenic (EPEC – causative agent of diarrhea in humans, rabbits, dogs, cats and horses); enteroinvasive (EIEC – found only in humans), verotoxigenic (VTEC – found in pigs, cattle, dogs and cats); enterohaemorrhagic (EHEC – found in humans, cattle, and goats, attacking porcine strains that colonize the gut in a manner similar to human EPEC strains) and enteroaggregative E. coli (EAggEC – found only in humans).

E. coli O157:H7 was first recognized as a pathogen as a result of an outbreak of unusual gastrointestinal illness in 1982. The outbreak was traced to contaminated hamburgers, and the illness was similar to other incidents in the United States and Japan. The etiologic agent of the illness was identified as a rare O157:H7 serotype of Escherichia coli in 1983. This serotype had only been isolated once before, from a sick patient in 1975.

Exceptional value for money
Rapid detection of all clinically relevant subtypes
Positive copy number standard curve for quantification
Highly specific detection profile
High priming efficiency
Broad dynamic detection range (>6 logs)
Sensitive to < 100 copies of target Accurate controls to confirm findings

Overview

• Columns bind proteins of interest while endotoxins flow through
• Reduce endotoxin levels to less than 0.01 EU/µg of protein
• Proteins are desalted
• Greater than 95% protein recovery
• Concentrate protein samples and remove detergents at the same time
• Effectively remove a wide range of detergents including SDS, Triton® X-100, CHAPS, NP-40, and Tween 20
• Purification is based on spin column chromatography that uses Norgen’s resin separation matrix

The ProteoSpin™ Total Protein Concentration, Detergent Clean-Up and Endotoxin Removal Kits are designed for the rapid removal of endotoxins from previously purified proteins or peptides, with protein recoveries of > 95% being achieved. Endotoxins, also known as lipopolysaccharides, are cell-membrane components of Gram-negative bacteria such as E. coli. Endotoxins liberated by Gram-negative bacteria are frequent contaminations of protein solutions derived from bioprocesses. Due to the high toxicity of endotoxins in vivo and in vitro, their removal from protein preparations is often necessary prior to the use of the protein in downstream applications. These kits efficiently reduce endotoxin levels to ≤ 0.01 EU/mg of protein, using spin column chromatography based on Norgen’s proprietary resin as the separation matrix. These kits are highly efficient in removing many different salts commonly used in the laboratory including, but not limited to, MgCl₂, NaCl, KCl, CaCl₂, LiCl and CsCl. The purified protein samples can be used in a number of downstream applications including sequencing, cloning, and in vitro and in vivo introduction into cells and organisms for various purposes. The simultaneous removal of salts while concentrating a dilute protein solution makes the kit a convenient method for preparing proteins before running many downstream applications such as SDS-PAGE, isoelectric focusing, X-ray crystallography, NMR spectroscopy, mass spectroscopy and other applications.

Mini Kit

The ProteoSpin™ Total Protein Concentration, Detergent Clean-Up and Endotoxin Removal Mini Kit is designed for the rapid removal of endotoxins from up to 200 μg of previously purified proteins or peptides, with protein recoveries of > 95% being achieved.

Maxi Kit

The ProteoSpin™ Total Protein Concentration, Detergent Clean-Up and Endotoxin Removal Maxi Kit is designed for the rapid removal of endotoxins from up to 4 mg of previously purified proteins or peptides, with protein recoveries of > 95% being achieved.

Details

Supporting Data

Figure 1 / 15

Previous

Next

Click for expanded view

Kit Specifications
Maximum Protein Input	200 μg
Maximum Column Volume Input	600 μL
Molecular Weight of Recovered Proteins	No Molecular Weight cut-off
Final Endotoxin Levels	≤ 0.01 EU/μg protein
Protein Recovery	90-95%
% Detergent Removal	90-95%
Detergents that can be Removed	Including Triton® X-100, CHAPS, NP-40 and Tween 20
Minimum Elution Volume	50 μL
Time to Complete 10 Purifications	20 minutes

Storage Conditions
All solutions should be kept tightly sealed and stored at room temperature. This kit is stable for 2 years from the date of shipment.

Component	Cat. 22800 (25 preps)	Cat. 22200 (4 preps)
Binding Buffer J	8 mL	8 mL
Binding Buffer N	4 mL	20 mL
Wash Solution M	50 mL	130 mL
Wash Solution CIP	20 mL	60 mL
Wash Solution N	30 mL	130 mL
Wash Solution NIP	20 mL	60 mL
Elution Buffer G	6 mL	20 mL
Endotoxin Removal Solution	1.5 mL	1.5 mL
Protein Neutralizer EF	2 mL	2 mL
Maxi Spin Columns (assembled with Collection Tubes)	–	4
Mini Spin Columns	25	–
Collection Tubes	25	–
Maxi Spin Columns (assembled with Collection Tubes)	4
Elution Tubes (1.7 mL)	25	–
Elution Tubes (50 mL)	4	–
Product Insert	1	1

Introduction

The HiPure Mini system provides a fast, simple, and cost-effective plasmid DNA miniprep method for routine molecular biology laboratory applications. HiPure Plasmid Mini Kits use silica membrane technology to eliminate the cumbersome steps associated with loose resins or slurries. Plasmid DNA purified with Mini Kits is immediately ready for use. Phenol extraction and ethanol precipitation are not required, and high quality plasmid DNA is eluted in a small volume of tris buffer or water.

Compared to other domestic products, Magen was the first to solve the stability problem of the column. Many other brands have unstable extraction concentrations, and the longer the time, the more unstable the column becomes. However, in our test of Magen kit, the quality and yield of plasmid extraction still remain stable after 5 years’ storage. For different customers, our kits can be customized. For example, Classic type is suitable for customers with low copy or unclear plasmid types. The rapid type is suitable for customers with high copy plasmids. Compared to many other brands, the plasmid DNA extracted by Magen has a longer preservation time and more thoroughly RNA removal.

Details

Specifications

Features	Specifications
Main Functions	Isolation up to 35μg plasmid DNA from 1-5ml bacterial culture
Applications	Enzyme digestion, sequencing, PCR, cloning, etc.
Purification method	Mini spin column
Purification technology	Silica technology
Process method	Manual (centrifugation or vacuum)
Sample type	Conventional plasmid, plasmid less than 30KB
Sample amount	High copy plasmid: 1-5ml culture mediumLow copy number plasmid : 5-10ml culture medium
Yield	5-35µg
Elution volume	≥30μl
Time per run	Complete 1-24 samples in 30 minutes
Liquid carrying volume per column	800µl
Binding yield of column	35µg

Principle

The HiPure Plasmid procedure is based on alkaline lysis of bacterial cells followed by adsorption of DNA onto silica in the presence of high salt. The unique silica membrane used in the kit completely replaces glass or silica slurries for plasmid DNA minipreps. The procedure consists of 3 basic steps: Preparation and clearing of a bacterial lysate by alkaline method，then transfer the supernatant to column to bind DNA. After washing proteins and other impurities, nucleic acid was finally eluted with low-salt buffer (10mm Tris, pH9.0, 0.5mm EDTA).

Advantages

• High purity – purified plasmid can be directly used in sequencing, enzyme digestion and PCR, etc.
• Fast – it takes only 1 minute to obtain supernatant by optimized solution (10 minutes for other brands)
• High yield – up to 35µg plasmid can be binded in one column

Kit Contents

Contents	P100102	P100103
Purification Times	100 Preps	250 Preps
RNase A	5 mg	10 mg
Buffer P1	30 ml	80 ml
Buffer P2	30 ml	80 ml
Buffer P3	40 ml	100 ml
Buffer PW1	60 ml	140 ml
Buffer PW2*	20 ml	50 ml
Elution Buffer	15 ml	30  ml
HiPure DNA Mini Columns II	100	250
2 ml Collection Tubes	100	250

Storage and Stability

The kit components can be stored dry at room temperature (15-25°C) and are stable for at least 18 months under these conditions. If any precipitates form in the buffers, warm at 37℃ to dissolve. After addition of RNase A，Buffer P1 is stable for 6 months when stored at 2-8°C.

Experiment Data

Purchase Guide

Name	CAT NO	Features	Sampleamount	Elutionvolume	Yield
HiPure Plasmid Mini Kit	P1001	Classic, Regular application	1-5ml LB	30-50μl	5–35µg/5ml
HiPure Plasmid Mini Kit II	P1002	Classic, Regular application	5-15ml LB	75-100μl	20-80µg/15ml
HiPure Plasmid Plus 96 Kit	P1006	Classic, Regular application	1-1.5ml LB	70-150μl	1-15μg/1ml
HiPure Fastfilter Plasmid Midi Kit	P1013	Fast	30-50ml LB	0.3-0.8ml	50–250µg/50ml
HiPure Fastfilter Plasmid Maxi Kit	P1014	Fast	100-200ml LB	0.5-1.5ml	0.4–1mg/200ml
Low endotoxin/Endotoxin-free Plasmid
HiPure Plasmid EF Maxi Kit	P1156	Classic method, General for all kinds of vectors	100-200ml LB	≥0.7ml	200–1500µg
HiPure Plasmid EF Mini Kit	P1154	Recommend for low copy vector, Thoroughly remove RNA	5-15ml LB	≥60μl	10–80µg
HiPure Plasmid EF 96 Kit	P1157	Recommend for low copy vector, Thoroughly remove RNA	1-5ml LBx96	≥75μl	30µg
HiPure Plasmid EF Midi Kit	P1231	Recommend for High copy vector, Low elution volume, High concentration	25-50ml LB	≥100μl	10-500μg
HiPure Plasmid EF Mega Kit	P1116	Recommend for High copy vector(>3μg/ml)	500ml LB	≥2ml	1-10mg

For any technical problems or customized products, please contact us.

F&Q about Endotoxin-free Plasmid Extraction Kit — P1156 ←click here

The HiPure Mini system provides a fast, simple, and cost-effective plasmid DNA miniprep method for routine molecular biology laboratory applications. HiPure Plasmid Mini Kits use silica membrane technology to eliminate the cumbersome steps associated with loose resins or slurries. Plasmid DNA purified with Mini Kits is immediately ready for use. Phenol extraction and ethanol precipitation are not required, and high quality plasmid DNA is eluted in a small volume of tris buffer or water.