4. The Second-dimensional SDS-PAGE (vertical) Protocol

Save this PDF as:
 WORD  PNG  TXT  JPG

Size: px
Start display at page:

Download "4. The Second-dimensional SDS-PAGE (vertical) Protocol"

Transcription

1 4. The Second-dimensional SDS-PAGE (vertical) Protocol I. PURPOSE This procedure outlines the steps that must be carried out in the seconddimension SDS-PAGE using vertical system. II. ENVIRONMENT All work must be done in a clean area. Special care should be taken to prevent contamination of the samples with keratin; i.e., wear gloves and Lab coat at all times when handling tubes, reagents and equipment that will come in contact with samples. This procedure should be completed under the temperature control which will improve gel-to-gel reproducibility. III. PROCESS SUMMARY The protocol includes preparation of the SDS-acrylamide gel, equilibration of the IPG strip(s) in equilibrium buffer (for reduction and alkylation), manipulation of the equilibrated IPG strip on SDS-gel, and running electrophoresis. IV. EQUIPMENT Name Description Special Requirements Adjustable Pipettes µl, µl Calibrated Pipette tips 200 and 1000µl Hoefer minive Gel size: 6x8 cm Amersham Hoefer SE 600 Gel size: 18x16 cm Amersham Ettan DALTtwelve System Gel size: 21.5x25 cm Amersham Ettan DALTsix System Gel size: 20.5x25.5 cm Amersham MultiTemp III system Cooling system (Hoefer SE 600) Amersham EPS 601 power supply Power supply (Hoefer SE 600) Amersham Lab glass ware For preparing solution Equilibration tube Amersham

2 V MATERIALS LIST Code No Name Supplier Milli-Q water In-house VW SDS (MW: g/mol) General Storage Bromophenol blue (BPB) Bio-Rad Low molecular weight std Amersham Agarose Amersham BP152-5 (5kg) Tris-Base (MW: g/mol) Fisher scientific BP318-5 (5kg) Glycine (MW: g/mol) Fisher scientific EC-890 ProtoGel National Diagnostics Acrylamide N,N - Acros organics Methylenebisacrylamide (37.5:1; 40% mix solution in water) Ammonium persulphate Amersham TEMED Bio-Rad PlusOne Dithiothreitol (DTT) Amersham Iodoacetamide 98% (IAM) Acros organics V. REAGENTS 1.5M Tris-Cl ph8.8 Tris (MW ) 182g Adjust to ph 8.8 and store at 4ºC 6 M HCI to ph 8.8 About 50ml Deionized water To 1000ml SDS equilibration buffer Final concentration 1.5M Tris-Cl ph mm Urea 6 M Make stock solution and aliquot 20 ml and Glycerol 30% stored at -20 o C. SDS 2% Bromophenol blue trace DTT 100 mg per 10 ml Add just prior to use Iodoacetamide 98% (IAM) 250 mg per 10 ml Add just prior to use Electrophoresis running buffer (10x) Tris-Cl 250 mm Stock this 10x electrophoresis running buffer Glycine 1.92 M at room temperature. Dilute it 10 times before using. SDS 1% 10% Ammonium persulfate solution (APS) Ammonium Persulfate 100mg Prepare fresh Deionized water 1ml 1% Agarose sealing solution Agarose 1g Heat the solution until the agarose is

3 Electrophoresis buffer 100ml completely dissolved Displacing solution (for DALT only) 1.5M Tris Cl ph8.8 25ml Should be made fresh; stored solution may Glycerol 50ml support microbial growth. Bromophenolblue 2mg Deionized water 25ml SDS 10% Overlay solution ( water saturated butanol) n or t-butanol 50 ml Shake and use top phase to overlay the Deionized water 5 ml monomer solutions VI. PROCEDURE NOTE: All equipment and materials that come into contact with gel samples must be clean and free of keratin contamination. Minimize handling and manipulations of samples. 1. Preparing SDS Gel: 1.1 Select the gel percentage: Single percentage offer better resolution for a particular MW window. A commonly used second-dimension gel for 2-D electrophoresis is a homogeneous gel containing 12.5% total acrylamide. The appropriate percentage gel is selected according to the range of separation desires. The table recommended acrylamide concentrations for protein separation: % Acrylamide in resolving gel Separation size range (MWx10-3 ) Single percentage: 5% % % % % Gradient: 5-15% % % Select the gel thickness: Either 1.0 mm or 1.5 mm

4 1.2 Casting homogeneous : SE 615 multiple caster can cast up 10. DALTtwelve caster can cast up 14. Be sure, that gel casters, spacers, separator sheets and cassettes or glass plates are perfectly clean and dry. Place the cassettes alternating with separator sheets into the caster. Block the spare space with blank cassettes when less than 12 (DALTtwelve caster) or less than 10 (SE 615 caster) should be prepared. Level the gel caster horizontally. 1.3 Preparing monomer solutions and casting homogeneous : 10 % 12.5% 15% Acrylamide stock 40%ig 25 ml 31 ml 37.5 ml 1.5M Tris-Cl ph ml 25 ml 25 ml 10% SDS 1 ml 1 ml 1 ml Water ml 36 ml TEMED 33µl 33µl 33µl 10% Ammonium persulfate 500µl 500µl 500µl Total volume 100 ml 100 ml 100 ml caster Hoefer minive Hoefer SE 600 Hoefer SE 615 Daltsix Dalttwelve Number of Acrylamide solution volume 850 ml 100 ml For 1.5 mm thick 60 ml per gel for 1.5mm thick 450 ml 500 ml for 1mm thick TEMED is added together with the stock solutions, it cannot catalyses the polymerization without ammonium persulfate. The ammonium persulfate should not be added before everything is in place, because there are only about 10 minutes left to pour the gel until it starts to gel. Pour the gel solution into the funnel using a 50 ml pipette, avoiding air bubbles. Stop pouring when the level has reached 1 cm below the upper edges of the casting cassettes.

5 Immediately pipette 1 ml overlay solution to each cassette. Let the gel polymerize for one hour (It is important to use the same volume for each cassette.). Inspect each gel cassette for eventual air bubbles. Gels with air bubbles should not be used. Wrapp the gel cassettes into wet paper towels and put it in a plastic bag. Store the usable in a cold room or refrigerator. 2. IPG strip equilibration 2.1 Take the equilibration buffer out of the freezer and prepare 1% agarose-solution. Heat the agarose solution on a heating stirrer until agarose gel is completely melted. The equilibration procedure also serves as reduction and alkylation for IPG strips. The table recommends suitable equilibration volume: IPG strip 13 or under 13cm 18cm 24 cm Equilibration buffer* 10ml 15ml 20ml * 40 ml aliquots are frozen and have to be split, add DTT and IDA always freshly Reduction Add DTT-containing equilibration buffer (100mg DTT/10ml buffer) to each IPG strip individual equilibration tube, place it on a rocker and equilibrate 15 min before pouring the buffer out Alkylation Add Iodoacetamide-containing equilibration buffer (250mg IAM/10ml buffer) to each IPG strip individual equilibration tube, place it on a rocker and equilibrate 15 min before pouring the buffer out. 3. Applying the IPG strip 3.1 Rinse the IPG strip with running buffer and place the strip with acidic end (+) to the left and plastic backing against one of the glass plates. With a thin plastic ruler, gently push the IPG strip down so that the entire lower edge of the IPG strip is in contact with the top surface of the slab gel. Ensure that no air bubbles are trapped between the IPG strip and the slab gel surfaces or between the gel backing and the glass plate. Note: If using 24 cm strip it has to be cut on the (-) end after placing against the glass plate and before pushing down. 3.2 Optional: If you want to apply a molecular weight marker proteins place sample application piece next to the acidic end.

6 3.3 Seal the IPG strip with 1% agarose gel. Allow the agarose to cool down until the tube can be held by fingers (60ºC) and then slowly pipette the amount required to seal the IPG strip in place. Pipetting slowly avoids introducing bubbles. 3.4 Load molecular weight marker proteins (Blue range + 10µl water) after inserting the precast gel cassettes into Hoefer 600 electrophoresis system and carefully pulling out the sample application piece. 4 SDS electrophoresis 4.1 Prepare electrophoresis running buffer and turn the cooling system on. The recommended temperature for SE 600 is 15ºC, for DALTsix is 5ºC and for DALTtwelve depends on running conditions. Electrophoresis buffer for Ettan DALTsix Gel thickness Anodic buffer (lower chamber) Volume [l] 1 mm 1xSDS electrophoresis buffer Cathodic buffer (upper chamber) 4.3 2xSDS electrophoresis buffer Volume [l] Preparing the electrophoresis unit DALTsix: Follow the instructions in Manual p12 DALTtwelve: Insert the precast gel cassettes. When the electrophoresis buffer has reached the desired temperature, insert loaded gel cassettes, push blank cassette inserts into any unoccupied slots. When 12 slots are occupied, the buffer level should be slightly below the level of the gaskets (0.5 cm). Note: Cassette insertion is aided by spraying the gel cassettes with a mist of SDS electrophoresis buffer prior to insertion.

7 4.3 Electrophoresis conditions: Electrophoresis system step Current (ma/gel) Power(w/gel) Duration(h:min) Hoefer minive 1.5 mm-thick : : mm-thick : :30 Hoefer SE mm-thick : : mm-thick : :00 DALTsix 1.0 mm-thick Phase (step) Temperature Power Duration (h:min) 1 5ºC (Yong) or 25 C 30W for 6 5 W per gel 30 min 600 V; 400 ma W for 6 7h 800 V; 800 ma (17) max 100 W per gel DALTtwelve 1.0 mm-thick Phase Temperature Power Duration (h:min) 1 28ºC 5 W per gel min W Max Approximately 5 h for ºC 5 W per gel min W max Approximately 6 h for ºC 5 W per gel min W max Approximately 8 h for ºC 5 W per gel min 2 80 W max Approximately h for ºC 1-2 W per gel min 2 Overnight The Table lists the recommended conditions for the Hoefer minive; SE600 and Ettan DALTsix and Ettan DALTtwelve systems.

8 Electrophoresis is performed at constant current (or constant power for DALTtwelve) in two steps. During the initial migration and stacking period, the current is approximately half of the value required for the subsequent separation. Do not cool SDS below 10ºC. For the smaller system (such as mini-gel), cooling is optional. The temperature control improves gel-to-gel reproducibility, especially if the ambient temperature of the laboratory fluctuates significantly. 4.4 Stop electrophoresis when the dye front line is approximately 1mm from the bottom of the gel. 4.4 After electrophoresis, remove from their gel cassettes in preparation for fixing and staining and cut a small piece of gel from the upper left corner (acidic side) off.

user manual electrophoresis system Ettan DALT II System second-dimension gel electrophoresis um /Rev. AO/09-00

user manual electrophoresis system Ettan DALT II System second-dimension gel electrophoresis um /Rev. AO/09-00 user manual electrophoresis system Ettan DALT II System second-dimension gel electrophoresis um 80-6476-53/Rev. AO/09-00 Page finder Ettan DALT II System....................................................................................1

More information

Week 1: Protein isolation and quantification

Week 1: Protein isolation and quantification Week 1: Protein isolation and quantification Objective The objective of this lab exercise is to obtain protein samples from fruit fly larvae, BCS and FBS, all of which are then quantitated in the preparation

More information

SERVAGel IEF 3-10 Precast Vertical Gels for Isoelectric Focusing

SERVAGel IEF 3-10 Precast Vertical Gels for Isoelectric Focusing INSTRUCTION MANUAL SERVAGel IEF 3-10 Precast Vertical Gels for Isoelectric Focusing (Cat. No. 43240, 43242) SERVA Electrophoresis GmbH Carl-Benz-Str. 7 D-69115 Heidelberg Phone +49-6221-138400, Fax +49-6221-1384010

More information

HiPer Immunoprecipitation Teaching Kit

HiPer Immunoprecipitation Teaching Kit HiPer Immunoprecipitation Teaching Kit Product Code: HTI016 Number of experiments that can be performed: 5 Duration of Experiment Storage Instructions The kit is stable for 6 months from the date of receipt

More information

Code Description Molecular Weight Separation Range

Code Description Molecular Weight Separation Range SPRINT NEXT GEL * A Ready-to-Pour Acrylamide Gel for the Rapid Electrophoresis of Proteins Code Description Molecular Weight Separation Range Size M312-100ML M312-500ML M311-100ML M311-500ML SPRINT NEXT

More information

GeneGel HyRes Starter Kit

GeneGel HyRes Starter Kit user manual DNA fragment analysis GeneGel HyRes Starter Kit for high-resolution electrophoretic DNA typing um 80-6452-59/Rev. B0/04-02 Page finder Introduction.............................. ii Preparing

More information

Guidelines for Using the Sage Science Pippin Pulse Electrophoresis Power Supply System

Guidelines for Using the Sage Science Pippin Pulse Electrophoresis Power Supply System Please note: the unsupported protocol described herein may not have been validated by Pacific Biosciences and is provided as-is and without any warranty. Use of this protocol is offered to those customers

More information

Lesson 3 Gel Electrophoresis of Amplified PCR Samples and Staining of Agarose Gels

Lesson 3 Gel Electrophoresis of Amplified PCR Samples and Staining of Agarose Gels Lesson 3 Gel Electrophoresis of Amplified PCR Samples and Staining of Agarose Gels What Are You Looking At? Before you analyze your PCR products, let s take a look at the target sequence being explored.

More information

TECHNICAL BULLETIN. Components System includes tank, lid, electrodes, electrophoresis and electroblotter modules, and accessories (see Table 1).

TECHNICAL BULLETIN. Components System includes tank, lid, electrodes, electrophoresis and electroblotter modules, and accessories (see Table 1). Sigma-Aldrich CVS10CBS omnipage mini dual Electrophoresis and Electroblotting System - Complete System for both Vertical Gel Electrophoresis and Electroblotting Catalog Number EP1301 Store at Room Temperature

More information

DGGE FOR Symbiodinium

DGGE FOR Symbiodinium DGGE FOR Symbiodinium By Camila Granados Cifuentes, IMaGeS Lab (Modified from Dr. Todd LaJeunesse and Dr. Juan Armando Sanchez BIOMMAR ) This protocol explains the step by step process for running a DGGE

More information

USDA RiceCAP DNA extraction using DNeasy Plant Mini Kit.

USDA RiceCAP DNA extraction using DNeasy Plant Mini Kit. DNA extraction using DNeasy Plant Mini Kit. Preparatory work: 1. If using the kit for the first time, add ethanol to buffer AW and buffer AP3/E to obtain the working solutions. 2. Preheat a water bath

More information

Principles and Practice of Agarose Gel Electrophoresis

Principles and Practice of Agarose Gel Electrophoresis Edvo-Kit #101 Principles and Practice of Agarose Gel Electrophoresis Experiment Objective: The objective of this experiment is to develop a basic understanding of electrophoretic theory, and to gain "hands-on"

More information

Agarose Gel Electrophoresis

Agarose Gel Electrophoresis Agarose Gel Electrophoresis Gel electrophoresis is a widely used technique for the analysis of nucleic acids and proteins. Agarose gel electrophoresis is routinely used for the preparation and analysis

More information

EXPERIMENT GENOMIC DNA ANALYSIS

EXPERIMENT GENOMIC DNA ANALYSIS EXPERIMENT GENOMIC DNA ANALYSIS Population diversity Studies We have 5 species of planarians (3 purchased from Carolina Biologicals, 2 obtained from the Levin lab) andmight have additional species found

More information

Protein Electrophoresis EZ-Run Protein Gel Solution EZ-Run Protein Standards EZ-Run Gel Staining Solution Traditional SDS-PAGE Reagents

Protein Electrophoresis EZ-Run Protein Gel Solution EZ-Run Protein Standards EZ-Run Gel Staining Solution Traditional SDS-PAGE Reagents Protein Electrophoresis EZ-Run Protein Gel Solution EZ-Run Protein Standards EZ-Run Gel Staining Solution Traditional SDS-PAGE Reagents Reliability. Purity. Certainty. Introduction Sodium dodecyl sulfate

More information

NOTE ACRYLAMIDE IS NEUROTOXIN YOU MUST WEAR GLOVES.

NOTE ACRYLAMIDE IS NEUROTOXIN YOU MUST WEAR GLOVES. GST Purfication and Pulldown Part I Instructor: David Deitcher TA: Kristy Lawton In order to study the function of a protein it is often useful to have that protein purified away from others in the cell.

More information

BIOLOGY 163 LABORATORY. RESTRICTION MAPPING OF PLASMID DNA (Revised Fall 2017)

BIOLOGY 163 LABORATORY. RESTRICTION MAPPING OF PLASMID DNA (Revised Fall 2017) BIOLOGY 163 LABORATORY RESTRICTION MAPPING OF PLASMID DNA (Revised Fall 2017) Physical mapping of genomes is an important part of modern molecular genetics. As it's name implies, physical mapping seeks

More information

Operation Manual. Concorde Vertical Gel Electrophoresis System. *This instrument is intended for laboratory use only.

Operation Manual. Concorde Vertical Gel Electrophoresis System. *This instrument is intended for laboratory use only. Concorde Vertical Gel Electrophoresis System Operation Manual Cat.no. R10-1031101 / R10-1031102 / R10-1031103 *This instrument is intended for laboratory use only http://www.recenttec.com E-mail : support@recenttec.com

More information

GE Healthcare. 2-D Electrophoresis. Principles and Methods

GE Healthcare. 2-D Electrophoresis. Principles and Methods GE Healthcare 2-D Electrophoresis Principles and Methods Introduction Introduction to this handbook This handbook is intended to provide guidelines for performing high-resolution two-dimensional (2-D)

More information

Frequent Difficulties With PFGE (Troubleshooting Tips)

Frequent Difficulties With PFGE (Troubleshooting Tips) Frequent Difficulties With PFGE (Troubleshooting Tips) 6 th PulseNet Latin America Meeting Buenos Aires, Argentina June 26 th 2008 Efrain M. Ribot, Ph.D. PulseNet Methods Development Laboratory Centers

More information

Multiphor II Electrophoresis System

Multiphor II Electrophoresis System data file Electrophoresis Multiphor II Electrophoresis System One system for 1-D or 2-D protein electrophoresis or DNA fragment analysis Introduction Multiphor II Electrophoresis System and accessories

More information

Positively Charged Membrane

Positively Charged Membrane BIOBOND NYLON MEMBRANES ProductInformation Technical Bulletin No. MB-570 June 1999 Size Quantity Positively Charged Membrane Neutral Membrane 30 cm x 3.5 m 1 roll N4781 N1031 30 cm x 12 m 1 roll N4906

More information

RFLP ANALYSIS OF DNA LABORATORY

RFLP ANALYSIS OF DNA LABORATORY RFLP ANALYSIS OF DNA LABORATORY BIG PICTURE You will be working with an essential research method widely used in genetics, conservation biology, and forensics. The lab is divided into three sections. Part

More information

Human DNA Alu Amplification by Polymerase Chain Reaction (PCR)* Laboratory Procedure

Human DNA Alu Amplification by Polymerase Chain Reaction (PCR)* Laboratory Procedure Human DNA Alu Amplification by Polymerase Chain Reaction (PCR)* Laboratory Procedure *Polymerase Chain Reaction is covered by patents owned by Hoffmann-La Roche, Inc. This experiment was adapted from Laboratory

More information

Instant-Bands Protein Sample Loading Buffer for SDS-PAGE. User s Manual. View Protein Bands in an SDS Gel. Instantly. EZBiolab.

Instant-Bands Protein Sample Loading Buffer for SDS-PAGE. User s Manual. View Protein Bands in an SDS Gel. Instantly. EZBiolab. Instant-Bands Protein Sample Loading Buffer for SDS-PAGE User s Manual View Protein Bands in an SDS Gel Instantly www.ezbiolab.com 2 Instant-Bands User s Manual Table of Contents Introduction 3 Storage

More information

Lambda (λ) DNA Restriction Digest and Electrophoresis Lab

Lambda (λ) DNA Restriction Digest and Electrophoresis Lab Lambda (λ) DNA Restriction Digest and Electrophoresis Lab Procedure DAY ONE: restriction digestion Today we will be exposing the lambda DNA to restriction enzymes. For background knowledge, make sure you

More information

Gel Electrophoresis and Analysis

Gel Electrophoresis and Analysis 6/28/2016 Gel Electrophoresis and Analysis B3 Summer Science Camp at Olympic High School Dr. Jennifer Weller Lab Method: Gel electrophoresis 2 6/28/2016 Electrophoresis: separating molecules in a charged

More information

SERVAGel SDS PAGE Starter Kit Precast Vertical Gels for Electrophoresis

SERVAGel SDS PAGE Starter Kit Precast Vertical Gels for Electrophoresis INSTRUCTION MANUAL SERVAGel SDS PAGE Starter Kit Precast Vertical Gels for Electrophoresis (Cat. No. 43200.01) SERVA Electrophoresis GmbH Carl-Benz-Str. 7 D-69115 Heidelberg Phone +49-6221-138400, Fax

More information

DNA RESTRICTION ANALYSIS

DNA RESTRICTION ANALYSIS DNA RESTRICTION ANALYSIS In this experiment, DNA from the bacteriophage Lambda (48,502 base pairs in length) is cut with a variety of restriction enzymes and the resulting fragments are separated using

More information

Code: IDK-006 Ver: 1 MTHFR A1298C. Detection system of the A1298C mutation in the human Methylen-tetraydrofolate reductase gene. Reg.

Code: IDK-006 Ver: 1 MTHFR A1298C. Detection system of the A1298C mutation in the human Methylen-tetraydrofolate reductase gene. Reg. Detection system of the A1298C mutation in the human Methylen-tetraydrofolate reductase gene Reg. MSP 21199 Valdense 3616, Montevideo 11700, Uruguay Phone (598) 2 336 83 01 Fax (598) 2 336 71 60 info@atgen.com.uy

More information

GE Healthcare. 2-D Electrophoresis. Principles and Methods

GE Healthcare. 2-D Electrophoresis. Principles and Methods GE Healthcare 2-D Electrophoresis Principles and Methods Handbooks from GE Healthcare Antibody Purification Handbook 18-1037-46 The Recombinant Protein Handbook Protein Amplification and Simple Purification

More information

qpcr Kit, DNA-free Product components 100 rxn 250 rxn Product description

qpcr Kit, DNA-free Product components 100 rxn 250 rxn Product description qpcr Kit, DNA-free For the PCR detection and identification of bacterial and fungal DNA using custom primers Product code A8514 Product components 100 rxn 250 rxn A 2.5x mastermix (3 mm MgCl 2 final concentration)

More information

TeloTAGGG Telomere Length Assay

TeloTAGGG Telomere Length Assay TeloTAGGG Telomere Length Assay Test Principle The test principle is shown schematically in the following figure: Note: The volumes used in the following are based on one experiment using a membrane size

More information

AGAROSE GEL ELECTROPHORESIS. Assiut University

AGAROSE GEL ELECTROPHORESIS. Assiut University AGAROSE GEL ELECTROPHORESIS By Prof. Dr. Asmaa Hussein Prof. of Zoonoses & Director of the MBRU Assiut University The standard method used to separate, identify electrophoresis and purify DNA fragments

More information

Amersham Western blotting

Amersham Western blotting Amersham Western blotting Selection guide www.gelifesciences.com Innovative solutions designed to meet your specific protein analysis needs Since 1990 when we first launched Amersham ECL we have continued

More information

Mini-V8 10 Vertical Gel Electrophoresis System

Mini-V8 10 Vertical Gel Electrophoresis System Instruction Manual Mini-V8 10 Vertical Gel Electrophoresis System CAT. SERIES 21078 Table of Contents 1. Notices to Customer... 1 1.1 Important Information... 1 1.2 Warnings... 1 2. Overview... 3 2.1 Description...

More information

RAINBOW GELS: AN INTRODUCTION TO ELECTROPHORESIS. STANDARDS 3.1.7, , Westminster College 3.3.7, , 3.3.

RAINBOW GELS: AN INTRODUCTION TO ELECTROPHORESIS. STANDARDS 3.1.7, , Westminster College 3.3.7, , 3.3. RAINBOW GELS: AN INTRODUCTION TO ELECTROPHORESIS STANDARDS 3.1.7, 3.1.10, 3.1.12 Westminster College 3.3.7, 3.3.10, 3.3.12 INTRODUCTION This laboratory will demonstrate the basics of electrophoresis and

More information

Revised 27 Aug rm (Vers. 4.1)

Revised 27 Aug rm (Vers. 4.1) Please use only the valid version of the package insert provided with the kit. INTENDED USE Vitamin Folic Acid Test is a microtiter plate test kit based on a microbiological assay which measures the total

More information

electrophoresis tech Versatile Separation Capabilities of the PROTEAN i12 IEF System

electrophoresis tech Versatile Separation Capabilities of the PROTEAN i12 IEF System electrophoresis tech note 6139 Versatile Separation Capabilities of the PROTEAN i12 IEF System ChengJun Sun, Adriana Harbers, Tom Berkelman, and Katy McGirr, Life Science Group, Bio-Rad Laboratories Inc.,

More information

Agarose Gel Electrophoresis Lab

Agarose Gel Electrophoresis Lab Agarose Gel Electrophoresis ACTIVITY AT A GLANCE Goal: This lab will determine the presence or absence of PCR products and uantify the size (length of the DNA molecule) of the products. Learning Objectives:

More information

Orexin A (HUMAN, MOUSE, RAT, PORCINE, OVINE,

Orexin A (HUMAN, MOUSE, RAT, PORCINE, OVINE, Orexin A (HUMAN, MOUSE, RAT, PORCINE, OVINE, BOVINE) Western Blot Kit Protocol (Catalog #WBK-003-30) PHOENIX PHARMACEUTICALS, INC. TABLE OF CONTENTS 1. Kit Contents...2 2. Storage...2 3. Introduction...3

More information

How to Set Up and Run Gel Electrophoresis

How to Set Up and Run Gel Electrophoresis How to Set Up and Run Gel Electrophoresis 2 Introduction Gel electrophoresis is a process by which one can separate and analyze various macromolecules based on their size and charge. The process is most

More information

Product Name : Simple mirna Detection Kit

Product Name : Simple mirna Detection Kit Product Name : Simple mirna Detection Kit Code No. : DS700 This product is for research use only Kit Contents This kit provides sufficient reagents to perform 20 reactions for detecting microrna. Components

More information

Long Ranger 50% Gel Solutions

Long Ranger 50% Gel Solutions Lonza Rockland, Inc. www.lonza.com scientific.support@lonza.com Scientific Support: 800-521-0390 Customer Service: 800-638-8174 Document # 18693-0311-06 Rockland, ME 04841 USA Long Ranger 50% Gel Solutions

More information

Southern Blot Protocol with Digoxigenin (DIG) probe JAX Mice strain: C57BL/6J Tg(C9orf72_i3)112Lutzy/J

Southern Blot Protocol with Digoxigenin (DIG) probe JAX Mice strain: C57BL/6J Tg(C9orf72_i3)112Lutzy/J Southern Blot Protocol with Digoxigenin (DIG) probe JAX Mice strain: 023099 C57BL/6J Tg(C9orf72_i3)112Lutzy/J Tail Genomic DNA extraction with QIAGEN DNeasy Blood & Tissue Kit (QIAGEN, cat. no.69504 or

More information

Experiment 3: The Chromatography of Organic Compounds

Experiment 3: The Chromatography of Organic Compounds Experiment 3: The Chromatography of Organic Compounds INTRODUCTION Very often, in an organic synthesis, a reaction will proceed to produce multiple products or perhaps will only partially form the desired

More information

Taura Syndrome Virus (TSV) RT-PCR Kit

Taura Syndrome Virus (TSV) RT-PCR Kit Revision No.: ZJ0001 Issue Date: Aug 28th, 2007 Taura Syndrome Virus (TSV) RT-PCR Kit Cat. No.: AR-0200-03 For use with Conventional PCR Instrument or Real time PCR Instrument User Manual For in vitro

More information

E.Z.N.A. Bacterial RNA Kit. R preps R preps

E.Z.N.A. Bacterial RNA Kit. R preps R preps E.Z.N.A. Bacterial RNA Kit R6950-00 5 preps R6950-01 50 preps July 2017 E.Z.N.A. Bacterial RNA Kit Table of Contents Introduction and Overview...2 Kit Contents/Storage and Stability...3 Before Beginning...4

More information

Minigel Protein Electrophoresis System Model P82 Operating and Maintenance Manual Rev. 0

Minigel Protein Electrophoresis System Model P82 Operating and Maintenance Manual Rev. 0 Visit us online to register your warranty www.thermoscientific.com/warranty Minigel Protein Electrophoresis System Model P82 Operating and Maintenance Manual 7007342 Rev. 0 Preface MANUAL NUMBER 7007342

More information

Gel Electrophoresis of Proteins

Gel Electrophoresis of Proteins Gel Electrophoresis of Proteins Comparative Proteomics: Protein Profiles of Fish Muscle Tissue Adapted by Audrey Dell Hammerich, 11/12/2016 Electrophoresis is the migration of charged molecules in a strong

More information

S-45. What Size Are Your Genes? Edvo-Kit #S-45. Experiment Objective: See page 3 for storage instructions.

S-45. What Size Are Your Genes? Edvo-Kit #S-45. Experiment Objective: See page 3 for storage instructions. Edvo-Kit #S-45 What Size Are Your Genes? S-45 Experiment Objective: The objective of this experiment is to develop an understanding that genetic mutations are inherited from one or both parents. Mutations

More information

KPL SignaLOCK ChemiWestern Kits (Film and Imager Analysis)

KPL SignaLOCK ChemiWestern Kits (Film and Imager Analysis) KPL SignaLOCK ChemiWestern Kits (Film and Imager Analysis) SignaLOCK HRP ChemiWestern Kit (Film) Catalog No. 54-53-00 SignaLOCK HRP ChemiWestern Kit (Imager) Catalog No. 54-54-00 SignaLOCK AP ChemiWestern

More information

UltraClean Midi Plasmid Prep Kit

UltraClean Midi Plasmid Prep Kit UltraClean Midi Plasmid Prep Kit Catalog No. Quantity 12700-20 20 Preps Instruction Manual Please recycle Version: 05232014 1 Table of Contents Introduction... 3 Protocol Overview... 3 Flow Chart... 4

More information

Human Myostatin, ELISA Kit (MSTN)

Human Myostatin, ELISA Kit (MSTN) Human Myostatin, ELISA Kit (MSTN) 96 Tests Catalog Number: MBS733837 Store all reagents at 2-8 C Valid Period: six months For samples: Cell culture fluid & body fluid & tissue homogenate Serum or blood

More information

Complete kit for the systematic analysis of protein kinases from a proteome such as cell lysate, tissue or microorganisms.

Complete kit for the systematic analysis of protein kinases from a proteome such as cell lysate, tissue or microorganisms. Protein Kinase Landscape (Large Format PEP) Catalog # AB000504 Complete kit for the systematic analysis of protein kinases from a proteome such as cell lysate, tissue or microorganisms. Please read this

More information

GeNei TM Gel Extraction Teaching Kit Manual

GeNei TM Gel Extraction Teaching Kit Manual Teaching Kit Manual Cat No. New Cat No. KT43 106279 KT43A 106300 KT43B 106301 Revision No.: 00280507 CONTENTS Page No. Objective 3 Principle 3 Kit Description 5 Materials Provided 7 Procedure 8 Observation

More information

Neural Stem Cells (ipsc from Blood Cells; Male)

Neural Stem Cells (ipsc from Blood Cells; Male) Applied StemCell, Inc. (866) 497-4180 www.appliedstemcell.com Datasheet Neural Stem Cells (ipsc from Blood Cells; Male) Product Information Catalog Number ASE-9234 (Male) Description Applied StemCell's

More information

NPTII ELISA Qualitative User Guide neomycin phosphotransferase II Catalog number: PSP 73000

NPTII ELISA Qualitative User Guide neomycin phosphotransferase II Catalog number: PSP 73000 List of contents Lot number Item 96 wells 288 wells 480 wells 4800 wells Antibody-coated 96-well microtiter plates 1 3 5 50 Peroxidase enzyme conjugate (bottles A and B) 0.150 ml 0.350 ml 0.550 ml 5.0

More information

QIAfilter Plasmid Midi Kit (Cat #: 12243)

QIAfilter Plasmid Midi Kit (Cat #: 12243) QIAfilter Plasmid Midi Kit (Cat #: 12243) Things to do before starting Add the provided RNase A solution to Buffer P1 before use. Use one vial of RNase A (centrifuge briefly before use) per bottle of Buffer

More information

RayBio Rat IL-6 ELISA Kit (For Lysates)

RayBio Rat IL-6 ELISA Kit (For Lysates) RayBio Rat IL-6 ELISA Kit (For Lysates) Catalog #: ELR-IL6-CL User Manual Last revised April 15, 2016 Caution: Extraordinarily useful information enclosed ISO 13485 Certified 3607 Parkway Lane, Suite 100

More information

The Biotechnology Education Company. Western Blot Analysis. Storage: See Page 3 for specific storage instructions EXPERIMENT OBJECTIVE:

The Biotechnology Education Company. Western Blot Analysis. Storage: See Page 3 for specific storage instructions EXPERIMENT OBJECTIVE: The Biotechnology Education Company EDVO-Kit # 317 Storage: See Page 3 for specific storage instructions EXPERIMENT OBJECTIVE: The objective of the experiment is for students to understand the theory and

More information

Geneaid DNA Isolation Kit (Yeast)

Geneaid DNA Isolation Kit (Yeast) Geneaid DNA Isolation Kit (Yeast) GEY100, GEY300 Advantages Sample: up to 2 10 8 yeast and other fungus species Yield: high yield, high quality DNA (A260/A280 = 1.8-2.0) Format: scalable DNA precipitation

More information

1 ml gel corresponds to ml of 75% (v/v) Glutathione Agarose suspension.

1 ml gel corresponds to ml of 75% (v/v) Glutathione Agarose suspension. 1 AFFINITY GST PURIFICATION Procedure for Use Glutathione Agarose 4 Resin DESCRIPTION Glutathione Agarose Resin is used to purify recombinant derivatives of glutathione S-transferases or glutathione binding

More information

Experiment 2: The Chromatography of Organic Compounds

Experiment 2: The Chromatography of Organic Compounds Experiment 2: The Chromatography of Organic Compounds INTRODUCTION When performing an organic reaction, it is very common to observe the formation of other compounds in addition to your desired product;

More information

E.Z.N.A. Tissue RNA Kit. R preps R preps

E.Z.N.A. Tissue RNA Kit. R preps R preps E.Z.N.A. Tissue RNA Kit R6688-00 5 preps R6688-01 50 preps May 2015 E.Z.N.A. Tissue RNA Kit Table of Contents Introduction and Overview...2 Kit Contents/Storage and Stability...3 Important Notes...4 Homogenization

More information

RayBio Apoptotic DNA Ladder Extraction Kit

RayBio Apoptotic DNA Ladder Extraction Kit RayBio Apoptotic DNA Ladder Extraction Kit User Manual Version 1.1 March 1, 2016 RayBio Apoptotic DNA Ladder Extraction (Cat#: 68SO-DNAL-S50) RayBiotech, Inc. We Provide You With Excellent Support And

More information

GUS Assays. 1. Label all tubes. Prepare solutions and have ready at hand.

GUS Assays. 1. Label all tubes. Prepare solutions and have ready at hand. GUS Assays I. Protein isolation A. Method for ~1g or more of tissue. 1. Label all tubes. Prepare solutions and have ready at hand. 2. Remove the tissue from the 80 o C freezer and thaw on ice. If the tissue

More information

Protocol for DNA extraction from FFPE Samples

Protocol for DNA extraction from FFPE Samples 25, ave Georges Lemaître - B-6041 Gosselies (Belgique) Tel : +32 (0)71 37 85 27 - Fax : +32 (0)71 34 78 79 Protocol for DNA extraction from FFPE Samples Step 1. Paraffin Removal 1 Equilibrate a heat block

More information

3. Close the bottom end of the column and apply the packing device on top. Pump water through the upper adaptor to remove air.

3. Close the bottom end of the column and apply the packing device on top. Pump water through the upper adaptor to remove air. INSTRUCTIONS FOR USE WorkBeads Protein A Product name Pack size Article number WorkBeads Protein A Bulk Media 1.5 ml 40605001 Bulk Media 5 ml 40605002 Bulk Media 10 ml 40605003 Bulk Media 100 ml 40605004

More information

COLORIMETRIC SANDWICH ELISA KIT INSTRUCTION MANUAL

COLORIMETRIC SANDWICH ELISA KIT INSTRUCTION MANUAL Page 1 of 7 COLORIMETRIC SANDWICH ELISA KIT INSTRUCTION MANUAL This product is for research use ONLY and not for human or animal therapeutic or diagnostic use. Page 2 of 7 Contents Page 3 I. Supplied Materials:

More information

Human IgG ELISA Kit. Strip well format. Reagents for up to 96 tests

Human IgG ELISA Kit. Strip well format. Reagents for up to 96 tests Human IgG ELISA Kit Strip well format. Reagents for up to 96 tests Catalog No. CS222A Quantity: 1 x 96 tests CS222B 5 x 96 tests Intended Use: Background: Assay Principle: This human immunoglobulin G antigen

More information

AFFINITY HIS-TAG PURIFICATION

AFFINITY HIS-TAG PURIFICATION DESCRIPTION Resins are products that allow batch or column purifications. This product is supplied as a suspension in 50% aqueous suspension containing 30 vol % ethanol. INSTRUCTIONS The resins are adapted

More information

Note: for laboratory research use only. RNA High-purity Total RNA Rapid Extraction Kit (Spin-column) Signalway Biotechnology

Note: for laboratory research use only. RNA High-purity Total RNA Rapid Extraction Kit (Spin-column) Signalway Biotechnology Note: for laboratory research use only RNA High-purity Total RNA Rapid Extraction Kit (Spin-column) Cat. #: RP1202 (50preps) Signalway Biotechnology I. Kit Content, Storage Condition and Stability Content

More information

Experion RNA StdSens Analysis Kit Instruction Manual

Experion RNA StdSens Analysis Kit Instruction Manual Experion RNA StdSens Analysis Kit Instruction Manual For Technical Service Call Your Local Bio-Rad Office or in the U.S. Call 1-800 4BIORAD (1-800-424-6723) Table of Contents Section 1 Introduction...1

More information

HiPer Random Amplification of Polymorphic DNA (RAPD) Teaching Kit

HiPer Random Amplification of Polymorphic DNA (RAPD) Teaching Kit HiPer Random Amplification of Polymorphic DNA (RAPD) Teaching Kit Product Code: HTBM031 Number of experiments that can be performed: 5 Duration of Experiment: Protocol: 3.5 hours Agarose Gel Electrophoresis:

More information

MOLECULAR GENETICS: TRANSFORMATION AND CLONING adapted by Dr. D. L. Vogelien

MOLECULAR GENETICS: TRANSFORMATION AND CLONING adapted by Dr. D. L. Vogelien Introduction MOLECULAR GENETICS: TRANSFORMATION AND CLONING adapted by Dr. D. L. Vogelien The field of molecular genetics has resulted in a number of practical applications that have been of tremendous

More information

minipcr TM Genes in Space Food Safety Lab: Mars Colony at Risk!

minipcr TM Genes in Space Food Safety Lab: Mars Colony at Risk! minipcr TM Genes in Space Food Safety Lab: Mars Colony at Risk! An E. coli outbreak affects astronaut food aboard the International Space Station. DNA samples from two food racks are analyzed to determine

More information

RayBio Human Caspase-3 ELISA Kit

RayBio Human Caspase-3 ELISA Kit RayBio Human Caspase-3 ELISA Kit Catalog #: ELH-CASP3 User Manual Last revised July 21, 2017 Caution: Extraordinarily useful information enclosed ISO 13485 Certified 3607 Parkway Lane, Suite 100 Norcross,

More information

Amersham * ECL * Gel horizontal electrophoresis system

Amersham * ECL * Gel horizontal electrophoresis system GE Healthcare Life Sciences Data file 28-9970-20 AB Electrophoresis products Amersham * ECL * Gel horizontal electrophoresis system Amersham ECL Gel and Amersham ECL Gel Box constitute a horizontal mini-gel

More information

RayBio Custom ELISA Kit

RayBio Custom ELISA Kit RayBio Custom ELISA Kit Catalog #: EL-PRELIM User Manual Last revised April 15, 2016 Caution: Extraordinarily useful information enclosed ISO 13485 Certified 3607 Parkway Lane, Suite 100 Norcross, GA 30092

More information

Human IgG ELISA Quantitation Set

Human IgG ELISA Quantitation Set Human IgG ELISA Quantitation Set Cat. No. E80-104 Components Supplied Affinity purified Goat anti-human IgG-Fc Coating Antibody A80-104A, 1 ml at 1 mg/ml Human Reference Serum, RS10-110-4, 0.1 ml HRP Conjugated

More information

Science Fair Project. Keep Calm and Split DNA Charisma Ware - Carleen McNees - Syracuse Junior High

Science Fair Project. Keep Calm and Split DNA Charisma Ware - Carleen McNees - Syracuse Junior High Science Fair Project Keep Calm and Split DNA Charisma Ware - Carleen McNees - Syracuse Junior High Problem How can you separate DNA when they are to small to see, let alone cut? Hypothesis If I put a group

More information

Human Angiotensin 2 (Ang2) ELISA

Human Angiotensin 2 (Ang2) ELISA Human Angiotensin 2 (Ang2) ELISA For the quantitative determination of human Ang2 in serum, plasma, cell culture fluid and other biological fluids Cat. No. KT-52748 For Research Use Only. Not for use in

More information

HiPer Gel Extraction Teaching Kit (Column Based)

HiPer Gel Extraction Teaching Kit (Column Based) HiPer Gel Extraction Teaching Kit (Column Based) Product Code: HTBM010 Number of experiments that can be performed: 10 Duration of Experiment Agarose Gel Electrophoresis: 1 hour Protocol: 1 hour Agarose

More information

Purification of DNA Oligonucleotides

Purification of DNA Oligonucleotides Purification of DNA Oligonucleotides Polyacrylamide gel: In a beaker, add 30 ml 30% acrylamide solution (To make this solution, add 29 g acrylamide to 1 g bis-acrylamide, bring to 100 ml with dh 2 O) 31.5

More information

Human immunoglobulin G(IgG) ELISA Kit

Human immunoglobulin G(IgG) ELISA Kit Human immunoglobulin G(IgG) ELISA Kit For the quantitative determination of human immunoglobulin G (IgG) concentrations in serum, plasma, cell culture supernates, urine, tissue homogenates, cell lysates.

More information

Let s Move It! Gel Electrophoresis Using Food Dye Student Guide

Let s Move It! Gel Electrophoresis Using Food Dye Student Guide Let s Move It! Gel Electrophoresis Using Food Dye Student Guide Purpose This lab explores the principle of electrophoresis, an important technique used in biochemistry and molecular biology. You will:

More information

Genome Edited ipscs APOE -/- Knockout

Genome Edited ipscs APOE -/- Knockout Applied StemCell, Inc. (866) 497-4180 www.appliedstemcell.com Datasheet Genome Edited ipscs APOE -/- Knockout Product Information Catalog Number Description Amount Parental Cell Line Gene Knockout Generated

More information

The preparation of native chromatin from cultured human cells.

The preparation of native chromatin from cultured human cells. Native chromatin immunoprecipitation protocol The preparation of native chromatin from cultured human cells. All solutions need to be ice cold. Sucrose containing solutions must be made up fresh on the

More information

ImmuLux Human IL-6 Fluorescent ELISA Kit

ImmuLux Human IL-6 Fluorescent ELISA Kit INSTRUCTIONS ImmuLux Human IL- Fluorescent ELISA Kit 3747 N. Meridian Road P.O. Box 7 Rockford, IL 05 840 53w Product Description Number Description 840 ImmuLux Human IL- Fluorescent ELISA Kit The Pierce

More information

Rat α-melanocyte stimulating hormone (α-msh) ELISA Kit

Rat α-melanocyte stimulating hormone (α-msh) ELISA Kit Rat α-melanocyte stimulating hormone (α-msh) ELISA Kit For the quantitative determination of rat α-melanocyte stimulating hormone (α-msh) concentrations in serum, plasma, tissue homogenates. This package

More information

Mouse Peptide YY (PYY) ELISA

Mouse Peptide YY (PYY) ELISA Mouse Peptide YY (PYY) ELISA For the quantitative determination of mouse PYY in serum, plasma, cell culture fluid and other biological fluids Cat. No. KT-58705 For Research Use Only. Not for use in diagnostic

More information

****** Competition ELISA Kit Instruction

****** Competition ELISA Kit Instruction FOR RESEARCH USE ONLY. NOT FOR DIAGNOSTIC PURPOSES ****** Competition ELISA Kit Instruction Kit name and catalog number Your analyte ELISA Kit, Catalog#: ***** Intended use The kit is used to detect the

More information

AnaTag HiLyte Fluor 647 Protein Labeling Kit

AnaTag HiLyte Fluor 647 Protein Labeling Kit AnaTag HiLyte Fluor 647 Protein Labeling Kit Catalog # 72049 Kit Size 3 Conjugation Reactions This kit is optimized to conjugate HiLyte Fluor 647 SE to proteins (e.g., IgG). It provides ample materials

More information

Speed Past the Twists and Turns in Protein Research

Speed Past the Twists and Turns in Protein Research BioResearch Speed Past the Twists and Turns in Protein Research Protein Electrophoresis Protein Electrophoresis from Lonza With over 50 years experience in the separation of molecular fragments, Lonza

More information

MATERIAL DATA SHEET. NOTE: Kit contains reagents sufficient for 10 x 30 μl reactions and 5 Western Blots (minigel. Reagents Provided in Kit

MATERIAL DATA SHEET. NOTE: Kit contains reagents sufficient for 10 x 30 μl reactions and 5 Western Blots (minigel. Reagents Provided in Kit Lot # XXXXX ITCH/AIP4 Ubiquitin Ligase Kit Cat. # K-270 MATERIAL DATA SHEET The mammalian Itchy homolog, or ITCH, (also known as Atrophin-1-interacting protein 4 or AIP4) is a HECT domain class ubiquitin

More information