Protein Sample Preparation and Analysis Application Manual - Analysis

Select a link below to access Protocols for Western Blotting.

• Western Blotting Using BioTrace™ NT Nitrocellulose Transfer Membrane (3.1.2)
• Western Blotting Using FluoroTrans® and BioTrace PVDF Membranes (3.1.3)

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 (Southerns, Northerns, Westerns).

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 membranes made from nitrocellulose, nylon, and PVDF for molecular detection applications.

Nitrocellulose, Nylon, and PVDF Membranes

Membrane Quick Selection Guide

Membrane Quick Selction Guide - Proteins

Membrane Quick Selection Guide - Nucleic Acids

Membranes and Devices for Use in Transfer and Immobilization Procedures

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

Product	Description	Binding Interaction	Method of Immobilization	Detection Methods
Biodyne® A  Membrane	Amphoteric Nylon 6,6	Hydrophobic & Electrostatic Baking	UV Crosslink Chemiluminescent	Radiolabeled Probes Enzyme-antibody Conjugates Chromogenic
Biodyne B  Membrane Biodyne Plus  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  Membrane	Negatively-charged Nylon 6,6	Electrostatic	Derivatization	Radiolabeled Probes Enzyme-antibody Conjugates Chromogenic
BioTrace™ NT  Membrane	100% Pure  Nitrocellulose	Hydrophobic &  Electrostatic	UV Crosslink Baking	Radiolabeled Probes Direct Stain, Fluorescence Enzyme-antibody Conjugates Chemiluminescent Chromogenic
BioTrace PVDF  Membrane	Polyvinylidene Fluoride	Hydrophobic	UV Crosslink Baking Alkaline Fixation	Direct Stain Enzyme-antibody Conjugates Chemiluminescent Chromogenic
FluoroTrans® W  Membrane	Polyvinylidene Fluoride	Hydrophobic	UV Crosslink Baking Alkaline Fixation	Direct Stain Enzyme-antibody Conjugates Chemiluminescent Chromogenic
UltraBind™  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 Membrane	100% Pure Nitrocellulose	Hydrophobic & Electrostatic	Incubation and Vacuum Filtration	Chemiluminescent Chromogenic Radiolabeled Probes Fluorescent

Membranes and Devices for Use in Transfer and Immobilization Procedures

Part 2 - Uses and Advantages of Membranes

Product	Works Best For	Also Suited FOr	Advantages
Biodyne® A  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  Membrane Biodyne Plus  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  Membrane	Reverse Dot Blots	Protein Immobilization, Affinity Purification, ELISAs	Negative charge over broad pH range Surface carboxyl groups can be derivatized
BioTrace™ NT  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  Membrane	Protein Transfers	Nucleic Acid Transfers, Protein Dot/Slot Blots	Chemical resistance No discoloration Nonflammable High strength
FluoroTrans® W  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™  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 Membrane	Dot Blots ELISAs	Protein Dot Blots	High binding capacity 96 well SBS-format Choice of housing colors

Select a link below to access Protocols for Affinity Activated or Activatable Membranes.

• Covalent Protein Immobilization Using UltraBind™ Affinity Membrane (3.2.2)
• ELISA Assays Using Immunodyne® ABC Membranes (3.2.3)
• ELISA and Immunoassay Using AcroWell™ 96 Multi-Well Filter Plates with BioTrace PVDF and NT Membranes (3.2.4)
• ELISA and Immunoassay Using AcroWell 96 Multi-Well Filter Plates with Chemiluminescent (DELFIA) Detection (3.2.5)
• Activation Protocol for Biodyne® C Membrane for Subsequent Covalent Linking of Ligands (3.2.6)

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 (Southerns, Northerns, Westerns).

Pall Corporation manufactures membranes made from nitrocellulose, nylon, and PVDF for molecular detection applications. Pall offers several activated membranes for covalent protein binding. Ultrabind modified polyethersulfone membrane has a high ratio of covalent to non-covalent protein binding. Immunodyne ABC membrane offers the sharp spot geometry of nylon with a proprietary covalent attachment chemistry.

See the Membrane Quick Selection Guide in the above section for further information.

See the table in the above section for Membranes and Devices for Use in Transfer and Immobilization Procedures.