b'The optimal product for IP, co-IP, and pull-down assays depends on the antibody or recombinant protein in the application (Table 6).Table 6. Choose your isolation strategy and find your product.Surface coating Choose this ifor ligand on theNonspecific Thermo Scientific and Invitrogen you use: magnetic beads Target binding IP protocol time Main benefits for IP productsProtein A, G, A/G,Primary antibodiesLow Dynabeads magneticDynabeads magnetic beads Dynabeads Protein Aor L from most species;beads: 40 min fastest, easiest protocol with lowDynabeads Protein A Immunoprecipitation Kitproteins A, G, andPierce beads: 130180 min nonspecific binding and high yieldDynabeads Protein GL bind differentand reproducibility Dynabeads Protein G Immunoprecipitation Kitantibody speciesPierce Magnetic IP-MS KitPierce Classic Magnetic IP/Co-IP Kit and subclasses with(Protein A/G) validated for massPierce Protein A/G Magnetic Beadsdifferent specificitiesspectrometry workflows Pierce Crosslink Magnetic IP/Co-IP KitPierce Magnetic IP-MS Kit (Protein A/G)Pierce Crosslink kit includes DSSPierce Protein L Magnetic BeadscrosslinkerSecondaryMouse IgG orLow Dynabeads magneticFast and easy protocolDynabeads M-280 Sheep AntiMouse IgGUnconjugatedantibodies rabbit IgG beads: 40 min Low nonspecific bindingDynabeads M-280 Sheep AntiRabbit IgGprimaryantibody Specific binding of mouse or rabbit IgGsEpoxy- and NHS- Any protein ligandUltralow Dynabeads magnetic beads: Covalent coupling of the Ab givesDynabeads Antibody Coupling Kitactivated beads (e.g., antibody,Ab coupling time:ultralow nonspecific bindingDynabeads Co-Immunoprecipitation Kitpeptide) overnightNo need for crosslinking Pierce NHS-activated magnetic beadsCo-IP protocol time:Gentle and efficient co-IP of even 3040 min large protein complexesPierce beads:Ab coupling time:30 60 minProtocol time: 120 minBiotinylatedStreptavidin Any biotinylatedLow Dynabeads magneticBinds any biotinylated proteinDynabeads MyOne Streptavidin T1antibody antibody or ligand beads: 40 min For samples high in soluble IgGsDynabeads M-280 StreptavidinPierce beads: 60120 min Pierce Streptavidin Magnetic BeadsRecombinant Ab lacking the FcPierce Magnetic IP-MS Kit (Streptavidin)regionRecombinantFusion tags Different beads bindLow Dynabeads His-tagPurify many different proteinsDynabeads His-Tag Isolation and Pulldownprotein proteins with thebeads: ~25 min incorporated with the same tag Pierce Anti-DYKDDDDK Magnetic Agarosefollowing tags: His,Pierce beads: ~70 min No need for antibodies Pierce HA-Tag Magnetic IP/Co-IP KitGST, DYKDDDDKPierce c-Myc-Tag Magnetic IP/Co-IP Kit(FLAG), HA, c-MycChoose these products if you use unconjugated primaryChoose these products if you use biotinylated antibodiesyour choice of antibody-binding productsantibodiesyour best choice when using a biotinylated depends on your downstream application, or if you do notantibody with streptavidin-coated beads for IP. These want the antibody co-eluted with your target protein. products are also useful in the following cases:Protein A, G, and A/G beads are most commonly usedIf you have a sample rich in soluble IgGsfor IP and co-IP applications since unconjugated primaryIf you have a recombinant antibody lacking Fc regionsantibodies that face towards the protein target bind directly to the coated beads in a short and simple incubation step.If you want to use streptavidin magnetic beads for Epoxy beads are often used to obtain ultralow nonspecificpull-down applications using a biotinylated protein as baitbinding, or to avoid crosslinking, since the antibody is covalently coupled to the beads and not eluted with theChoose these products if you have a recombinant target protein. protein with a fusion tagthe most popular fusion tags for The Invitrogen epoxy beads and co-IP kit (includingrecombinant protein expression are covered by Pierce and optimized buffers) are recommended for co-IP applicationsDynabeads products. DYKDDDDK (FLAG epitope), HA-tag, involving larger protein complexes. and c-Myc tags are ideal for IP/co-IP applications, while His-tag and GST-tagged proteins are for pull-down assays.36 Protein purification thermofisher.com/proteinbiology'