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Western Blotting

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Proteomics, Protein Chemistry, Protein Purification

Western Blotting Sections

Complete Protein Application Manual

Complete Protein Application Manual

Western Blotting Introduction

Membranes have been standard tools in molecular biology laboratories since the 1970s, when it was discovered that biomolecules could be spotted directly onto membranes (spot ELISA or DNA dot blots) or transferred from gels (Southern blots, Northern blots, Western blots).

The first membranes used for molecular detection were made from nitrocellulose, which has high affinity for biomolecules and low background with many detection techniques. PVDF membranes were found to have higher affinity for proteins than nitrocellulose and are now favored in Western transfers, although nitrocellulose remains in widespread use.

Pall Corporation manufactures Western blotting membranes made from nitrocellulose, nylon, and PVDF for molecular detection applications.

Table 3.1

Part 1 - Western Blotting Membranes Binding Interaction, Method of Immobilization, and Detection Methods


Product	Description	Binding Interaction	Method of Immobilization	Detection Methods
Biodyne® A Transfer Membrane	Amphoteric Nylon 6,6	Hydrophobic & Electrostatic Baking	UV Crosslink Chemiluminescent	Radiolabeled Probes Enzyme-antibody Conjugates Chromogenic
Biodyne B Transfer Membrane Biodyne Plus Transfer Membrane	Positively-charged Nylon 6,6	Electrostatic	Can be baked or UV crosslinked although not required	Radiolabeled Probes Enzyme-antibody Conjugates Chemiluminescent Chromogenic
Biodyne C Transfer Membrane	Negatively-charged Nylon 6,6	Electrostatic	Derivatization	Radiolabeled Probes Enzyme-antibody Conjugates Chromogenic
BioTrace™ NT Transfer Membrane	100% Pure Nitrocellulose	Hydrophobic & Electrostatic	UV Crosslink Baking	Radiolabeled Probes Direct Stain, Fluorescence Enzyme-antibody Conjugates Chemiluminescent Chromogenic
BioTrace PVDF Transfer Membrane	Polyvinylidene Fluoride	Hydrophobic	UV Crosslink Baking Alkaline Fixation	Direct Stain Enzyme-antibody Conjugates Chemiluminescent Chromogenic
FluoroTrans® W Western Transfer Membrane	Polyvinylidene Fluoride	Hydrophobic	UV Crosslink Baking Alkaline Fixation	Direct Stain Enzyme-antibody Conjugates Chemiluminescent Chromogenic
UltraBind™ Affinity Membrane	Modified Polyethersulfone	Covalent to Amine Groups	Air Drying	Radiolabeled Probes Enzyme-antibody Conjugates Chromogenic
Immunodyne® ABC Membrane	Modified Nylon 6,6	Covalent to Amine and Carboxyl Groups	Air Drying	Chemiluminescent Chromogenic Radiolabeled Probes Fluorescent
AcroWell™ Filter Plates with BioTrace PVDF Membrane	Polyvinylidene Fluoride	Hydrophobic	Incubation and Vacuum Filtration	Chemiluminescent Chromogenic Fluorescent
AcroWell Filter Plates with BioTrace NT Transfer Membrane	100% Pure Nitrocellulose	Hydrophobic & Electrostatic	Incubation and Vacuum Filtration	Chemiluminescent Chromogenic Radiolabeled Probes Fluorescent

Table 3.1

Part 2 - Uses and Advantages of Membranes


Product	Works Best For	Also Suited FOr	Advantages
Biodyne® A Transfer Membrane	Colony/Plaque Lifts, DNA and RNA Transfers	Gene Probe Assays, DNA Fingerprinting, Nucleic Acid Dot/Slot Blots, Replica Plating, ELISAs	High Sensitivity Low Background Net charge can be controlled by chanign pH
Biodyne B Transfer Membrane Biodyne Plus Transfer Membrane	DNA and RNA Transfers, Multiple Reprobings, Replica Plating	DNA Fingerprinting Nucleic Acid Dot/Slot Blots, Colony/Plaque Lifts (Biodyne B membrane)	Positive charge over broad pH range Highest sensitivity for nucleic acid applications (Biodyne B membrane) No need to bake or UV crosslink
Biodyne C Transfer Membrane	Reverse Dot Blots	Protein Immobilization, Affinity Purification, ELISAs	Negative charge over broad pH range Surface carboxyl groups can be derivatized
BioTrace™ NT Transfer Membrane	Colony/Plaque Lifts	Nucleic Acid and Protein Transfers, Protein Dot/Slot Blots	Excellent strength No support fabric No detergents added 100% pure nitrocellulose
BioTrace PVDF Transfer Membrane	Protein Transfers	Nucleic Acid Transfers, Protein Dot/Slot Blots	Chemical resistance No discoloration Nonflammable High strength
FluoroTrans® W Western Transfer Membrane	FluoroTrans W: Western Transfers FluoroTrans: N-terminal Protein Sequencing	Southern Transfers	Strong protein binding Sensitive detection Very low burn-through Good chemical compatibility
UltraBind™ Affinity Membrane	Solid-phase ELISAs	Affinity Chromatography, Hybridoma Screening	Permanent binding No preactivation required High protein binding capacity
Immunodyne® ABC Membrane	Immuno Assays Diagnostic Applications Reverse Dot Blots	Enzyme Biosensor Applications, ELISAs	Activated surface No preactivation required Intrinsically hydrophilic
AcroWell™ Filter Plates with BioTrace PVDF Membrane	Protein Detection ELISAs	Dot Blots, Protein:Protein Interactions	High binding capacity Chemical resistance 96 well SBS-format Choice of housing colors
AcroWell Filter Plates with BioTrace NT Transfer Membrane	Dot Blots ELISAs	Protein Dot Blots	High binding capacity 96 well SBS-format Choice of housing colors

Figure 3.1

Nitrocellulose, Nylon, and PVDF Western Blotting Membranes

Western Blotting Nitrocellulose, Nylon, and PVDF Membranes

Western Blotting Membrane Quick Selection Guide

Proteins

Western Blotting Membrane Selection Guide Proteins

Western Blotting Membrane Quick Selection Guide Continued

Nucleic Acids

Western Blotting

Western Blotting Sections

Complete Protein Application Manual

Technical Appendices

Introduction

Pre-analytical Applications

Protein Analysis

Purification Applications

Supporting Technologies

Western Blotting Introduction

Table 3.1

Table 3.1

Figure 3.1

Western Blotting Membrane Quick Selection Guide

Western Blotting Membrane Quick Selection Guide Continued