ProteoMiner Protein Enrichment Kits - Bio-Rad

1 ProteoMiner ProteinEnrichment KitsInstruction ManualCatalog #163-3003163-3006163-3007163-3008163-300 9163-3010163-3011163-3012 For Technical Support, contact your local Bio-Rad office, orin the US, call 1-800-4 BIORAD (1-800-424-6723)10010636 3/12/2009 7:51 AM Page 1 Table of ContentsSection 1 Introduction ..1 Section 2 Kit Specifications ..1 Section 3 Storage Conditions ..3 Section 4 Reagent Preparation ..4 Section 5 Sample Considerations ..4 Section 6 Instructions for Use With ProteoMiner Large-Capacity kits (Catalog #s 163-3007 and 163-3009).

2 4 Section 7 Instructions for Use With ProteoMinerSmall-Capacity kits (Catalog #s 163-3006 and 163-3008) ..7 Section 8 Instructions for Use With SequentialElution Large-Capacity Kit (Catalog #163-3011) ..9 Section 9 Instructions for Use With SequentialElution Small-Capacity Kit(Catalog #163-3010) ..12 Section 10 Instructions for Use With Bulk Beads(Catalog #163-3012) ..15 Section 11 Appendix ..17 Section 12 References ..18 Section 13 Product Information ..1810010636 3/12/2009 7:51 AM Page TOC1 Section 1 Introduction The ProteoMiner technology is a novel sample preparation tool used for thecompression of the dynamic range of the Protein concentration in complexbiological samples.

3 High-abundance proteins present in complex biologicalsamples like sera or plasma, make the detection of medium- and low-abundance proteins extremely challenging. This technology provides a methodof overcoming this challenge, allowing for the exploration of the entireproteome. This is accomplished through the use of a large, highly diverse bead-basedlibrary of combinatorial peptide ligands. When complex biological samples areapplied to the beads, the high-abundance proteins saturate their high affinityligands and excess Protein is washed away.

4 In contrast, the medium- and low-abundance proteins are concentrated on their specific affinity ligands. Thisreduces the dynamic range of Protein concentrations while maintainingrepresentatives of all proteins within the original 2 Kit SpecificationsProteoMiner Small-Capacity Kit (Catalog #163-3006)Provides reagents for processing 10 samples. Compatible with 2-D gelelectrophoresis and other downstream Protein separation analysis methods. Spin Columns. 10 columns each containing 500 l bead slurry (4% beads,20% v/v aqueous EtOH), 20 l settled bead volume Wash Buffer.

5 50 ml PBS buffer (150 mM NaCl, 10 mM NaH2PO4, pH ) Elution Reagent. 2 vials, lyophilized urea CHAPS (8 M urea, 2% CHAPS) Rehydration Reagent. 5 ml, 5% acetic acid Collection Tubes. 20 capless, 2 ml centrifuge tubes; 10 capped, 2 mlcentrifuge tubes Instruction Manual Quick Start GuideProteoMiner Large-Capacity Kit (Catalog #163-3007)Provides reagents for processing 10 samples. Compatible with 2-D gelelectrophoresis and other downstream Protein separation analysis methods. Spin Columns. 10 columns each containing 500 l bead slurry (20%beads, 20% v/v aqueous EtOH), 100 l settled bead volume110010636 3/12/2009 7:51 AM Page 1 Wash Buffer.

6 50 ml PBS buffer (150 mM NaCl, 10 mM NaH2PO4, pH ) Elution Reagent. 5 vials, lyophilized urea CHAPS (8 M urea, 2% CHAPS) Rehydration Reagent. 5 ml, 5% acetic acid Collection Tubes. 20 capless, 2 ml centrifuge tubes; 10 capped, 2 mlcentrifuge tubes Instruction Manual Quick Start GuideProteoMiner Introductory Small-Capacity Kit (Catalog #163-3008)Contains two spin columns, all other reagents are the same as in theProteoMiner small-capacity kit above. Introductory kit provides reagents forprocessing two Introductory Large-Capacity Kit (Catalog #163-3009)Contains two spin columns, all other reagents are the same as in theProteoMiner large-capacity kit above.

7 Introductory kit provides reagents forprocessing two Sequential Elution Small-Capacity Kit (Catalog #163-3010)This kit combines the ProteoMiner small-capacity kit (catalog #163-3006) andthe ProteoMiner sequential elution reagents (catalog #163-3003) and isdesigned to provide multiple elution options for researchers using SELDI orother downstream Protein separation analysis methods, and who wish toaccess additional proteins. This kit is NOT compatible with 2-D Sequential Elution Large-Capacity Kit (Catalog #163-3011)` This kit combines the ProteoMiner large-capacity kit (catalog #163-3007) andthe ProteoMiner sequential elution reagents (catalog #163-3003) and isdesigned to provide multiple elution options for researchers using SELDI orother downstream Protein separation analysis methods, and who wish toaccess additional proteins.

8 This kit is NOT compatible with 2-D Sequential Elution Reagents (Catalog #163-3003)To be used in combination with the ProteoMiner kits (catalog #s163-3006 or163-3007), this product provides reagents for processing 10 samples. Thesequential elution reagents are available for researchers using SELDI or otherdownstream Protein separation analysis methods, and who wish to access210010636 3/12/2009 7:51 AM Page 2additional proteins. These reagents are NOT compatible with 2-D gelelectrophoresis. Elution Reagent 1. 5 ml 1 M sodium chloride, 20 mM HEPES, pH Elution Reagent 2.

9 5 ml 200 mM glycine, pH Elution Reagent 3. 5 ml 60% ethylene glycol in water Elution Reagent 4. 5 ml 2-propanol, acetonitrile, acid Plasma Preparation Buffer. ml 1 M sodium citrate, 20 mM HEPES, pH Collection tubes. 30 capped, 2 ml centrifuge tubesItems required but not provided: Microcentrifuge or vacuum manifold (available through Bio-Rad , catalog#732-6470) Pipet Proteomics grade water (available through Bio-Rad , catalog #163-2091) ProteoMiner Beads (Catalog #163-3012)Provides 525 mg of dry beads, columns and reagents not 3 Storage ConditionsStore the unopened kit at 4 C.

10 After reconstituting lyophilized elution reagentstore any remaining material at 20 C for up to one week. 310010636 3/12/2009 7:51 AM Page 3 Section 4 Reagent PreparationPrepare elution reagent by adding 610 l rehydration reagent to one viallyophilized elution reagent. Following rehydration, each vial will contain enoughelution reagent for processing two samples. If preparing an uneven number ofcolumns, you will have remaining material that will be required for subsequentpreparations. Remaining material may be stored at 20 C for up to one , it is recommended to make this solution fresh each time you use thekit.