A vial of liquid magnetic beads for protein purification

Accelerate proteomics workflow throughput (HTS) using magnetic separations

Speed and sample purity are major concerns for proteomics researchers, and strategies to achieve higher throughput for sample-handling and data-processing are desired. Magnetic particles comprise a convenient solid support for a variety of assays and procedures based on affinity purification. Paramagnetic beads are especially well-suited for automated procedures because instrumentation exists to easily mix, incubate, and separate the particles in 96-well plates without columns or centrifugation. Such ease of manipulation enables rapid processing of many samples and seamless integration and compatibility with entire workflows.


Automation-verified Pierce Magnetic Beads

Pierce Magnetic Beads are small, 1 μm particles that allow for high-throughput screening, purification, and immunoprecipitation with ng–μg protein yields.  Alternatively, Pierce Magnetic Agarose consists of significantly larger, 10–40 μm particles and is used for high-capacity protein purification at mg–g scales. Both Pierce bead types can be automated on KingFisher Instruments or used in a manual format with magnetic stands.

Magnetic beadAffinity targetMagnetic bead kitsBinding strategyKingFisher software protocols
Anti-HAHA-tagged fusion proteinsHA-Tag Magnetic IP/Co-IP kitDirect
Anti-c-MycMyc-tagged fusion proteinsMyc-Tag Magnetic IP/Co-IP kitDirect
Ni-NTAHis-tagged fusion proteins Direct
NHS esterCovalent immobilization of primary amine molecules
(proteins, peptides, etc.) for affinity purification
Direct Magnetic IP kitDirect

IP protocol

Coupling protocol

Protein A/GAntibodies (IgG containing Fc)Classic Magnetic IP kit
Crosslink Magnetic IP kit
Indirect
Protein AAntibodies (IgG containing Fc) Indirect
Protein GAntibodies (IgG containing Fc) Indirect
Protein LAntibodies (Ig containing kappa light chains) Indirect
StreptavidinBiotin or desthiobiotin Indirect

Flex Instrument

Magnetic agarose beadAffinity targetBinding strategyKingFisher instrument protocols
Anti-DYKDDDDKDYKDDDDK-tagged fusion proteins 

Acid elution

Peptide elution

GlutathioneGlutathione S-transferase (GST) fusion proteins 
Ni-NTAHis-tagged fusion proteinsDirect
Ni-IMACHis-tagged fusion proteins, EDTA compatible

Direct

(EDTA compatible chemistry)

Protein A/GAntibodies (IgG containing Fc)Indirect
Magnetic beadAffinity targetMagnetic bead kitsBinding strategyKingFisher software protocols
Anti-HAHA-tagged fusion proteinsHA-Tag Magnetic IP/Co-IP kitDirect
Anti-c-MycMyc-tagged fusion proteinsMyc-Tag Magnetic IP/Co-IP kitDirect
Ni-NTAHis-tagged fusion proteins Direct
NHS esterCovalent immobilization of primary amine molecules
(proteins, peptides, etc.) for affinity purification
Direct Magnetic IP kitDirect

IP protocol

Coupling protocol

Protein A/GAntibodies (IgG containing Fc)Classic Magnetic IP kit
Crosslink Magnetic IP kit
Indirect
Protein AAntibodies (IgG containing Fc) Indirect
Protein GAntibodies (IgG containing Fc) Indirect
Protein LAntibodies (Ig containing kappa light chains) Indirect
StreptavidinBiotin or desthiobiotin Indirect

Flex Instrument

Magnetic agarose beadAffinity targetBinding strategyKingFisher instrument protocols
Anti-DYKDDDDKDYKDDDDK-tagged fusion proteins 

Acid elution

Peptide elution

GlutathioneGlutathione S-transferase (GST) fusion proteins 
Ni-NTAHis-tagged fusion proteinsDirect
Ni-IMACHis-tagged fusion proteins, EDTA compatible

Direct

(EDTA compatible chemistry)

Protein A/GAntibodies (IgG containing Fc)Indirect

KingFisher Magnetic Particle Processors

Prepare samples efficiently from a variety of materials with Thermo Scientific KingFisher Purification Systems, which offer highly versatile, automated magnetic-particle processing for DNA/RNA, protein, or cell purification from virtually any source. Using revolutionary magnetic particle separation technology, these systems provide excellent reproduciblity and quality. Thermo Scientific BindIt Software provides integrated or remote control of KingFisher Instruments. Ready-to-use BindIt Software protocols (.bdz files) are available for variety of applications with Pierce Magnetic Beads.

88845-88800-Protein-A-Mag

Figure 1. Highly reproducible IgG purification. Pierce Protein A Magnetic Beads and a KingFisher Instrument were used to purify IgG from rabbit serum in 16 wells of a 96 deep-well plate. Reducing SDS-PAGE results in separate heavy- and light-chain bands of purified IgG. Yield and purity are identical across all wells, confirming the high quality and reproducibility of the beads and instrument.

Figure 2. Consistency of "Pierce High-Capacity Ni-IMAC MagBeads, EDTA Compatible" purifications performed on the KingFisher Flex. Cell culture supernatant (Expi293) containing over-expressed His-tagged HSA (50 mg total protein) was applied to Pierce High-Capacity Ni-IMAC MagBeads, EDTA-Compatible (10ul settled beads) in a 96-well plate. Beads were processed using the KingFisher Flex. Binding was performed with all samples for 30 minutes. The beads were then washed twice, and bound protein was eluted with a 15-minute incubation in Elution Buffer. The eluates were resolved on an SDS-PAGE gel and stained with GelCode Blue (Cat. No. 24594). Gel lanes were normalized to equivalent volume.
M = MW marker, L = lysate load, 1-12 = corresponding elution from plate row. CV’s <10%

SDS-PAGE gel showing consistent performace of Pierce High-Capacity Ni-IMAC MabBeads in purifying proteins

Automated magnetic protein purification selection guide

 Magnetic beads
(ng to ug binding capacity)
Magnetic agarose beads
(mg capacity)
IP-MSProtein A/G
Streptavidin
 
IP/co-IPProtein A/G
Protein A
Protein G
 
Biotin IPStreptavidin 
ChIPProtein A/G
Protein A
Protein G
 
Epitope tagAnti-HA
Anti-c-Myc
Glutathione
Anti-DYKDDDDK
Antibody purificationProtein A/G
Protein A
Protein G
Protein L
Protein A/G
IMACNi-NTANi-NTA
Ni-IMAC (EDTA-compatible)
Surface reactive

NHS

 

Automated protein purification workflow

KingFisher Sample Preparation Systems