Shop All Immunoprecipitation & Pull-Down Kits

Pierce™ Streptavidin Coated Plate IP Kit (Thermo Scientific™)

The Thermo Scientific Pierce Streptavidin Coated Plate IP Kit uses coated streptavidin microplates to perform immunoprecipitation assays in 96-well plates without resins, beads, centrifugation or magnets.

Pierce Coated Plate IP Kits enable rapid immunoprecipitation of multiple samples without the usual tedium of pipetting, centrifuging and separating beaded affinity resin in individual microcentrifuge tubes. Immunoprecipitation is accomplished using coated 96-well microplates rather than beaded agarose resin. The plate format allows for faster processing of multiple samples.

Features of the Streptavidin Coated Plate IP Kit:

• Ready-to-use, high quality coated plates provide high capacity and consistency
• Plate format best suited for simultaneously processing multiple samples and their control conditions
• Faster, easier and more thorough washing than with traditional tube/resin IP methods
• Uses familiar and convenient ELISA tools (multichannel pipettors and plate washing); no tedious separation of supernatant from pelleted resin beads, and no tubes to open and close and centrifuge
• Coated plates are 96-well strip plates, convenient for experiments requiring only a partial plate
• Easy-to-follow instructions, including detailed explanation of appropriate controls

Applications:
• Rapid, trouble-free immunoprecipitation of multiple samples and their controls
• Immunoprecipitation when plate-based tools and techniques are preferred

Streptavidin is a protein that binds specifically and very strongly to biotin; therefore, the Streptavidin Coated Plate IP Kit (Part No. 45360) is appropriate for immunoprecipitation when using a biotin-labeled (biotinylated) antibody. In fact, this kit can be used to affinity purify a binding partner to any antibody species or subclass or any other protein or molecule that is biotinylated. Because the streptavidin-biotin affinity interaction is so strong, the elution step generally will dissociate only the antigen (binding partner), not the biotinylated antibody or "bait" protein.

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Pierce™ Protein A/G Coated Plate IP Kit

Pierce™ Protein A/G Coated Plate IP Kit (Thermo Scientific™)

The Thermo Scientific Pierce Protein A/G Coated Plate IP Kit uses coated Protein A/G microplates to perform immunoprecipitation assays in 96-well plates without resins, beads, centrifugation or magnets.

Pierce Coated Plate IP Kits enable rapid immunoprecipitation of multiple samples without the usual tedium of pipetting, centrifuging and separating beaded affinity resin in individual microcentrifuge tubes. Immunoprecipitation is accomplished using coated 96-well microplates rather than beaded agarose resin. The plate format allows for faster processing of multiple samples.

Features of the Protein A/G Coated Plate IP Kit:

• Ready-to-use, high quality coated plates provide high capacity and consistency
• Plate format best suited for simultaneously processing multiple samples and their control conditions
• Faster, easier and more thorough washing than with traditional tube/resin IP methods
• Uses familiar and convenient ELISA tools (multichannel pipettors and plate washing); no tedious separation of supernatant from pelleted resin beads, and no tubes to open and close and centrifuge
• Coated plates are 96-well strip plates, convenient for experiments requiring only a partial plate
• Easy-to-follow instructions, including detailed explanation of appropriate controls

Applications:
• Rapid, trouble-free immunoprecipitation of multiple samples and their controls
• Immunoprecipitation when plate-based tools and techniques are preferred

Protein A and Protein G are different proteins that bind to immunoglobulins (primarily only IgG). Typically, Protein A is preferred for use with Rabbit polyclonal antibodies, while Protein G is preferred for use with mouse antibodies (especially monoclonals of the IgG1 subclass). Protein A/G is a recombinant of Protein A and Protein G that has the additive binding properties of both proteins. Compare Protein A, Protein G and other antibody-binding proteins.

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Pierce™ Direct Magnetic IP/Co-IP Kit (Thermo Scientific™)

Thermo Scientific Pierce Direct Magnetic IP/Co-IP Kit uses NHS-chemistry to covalently immobilize IP antibodies onto magnetic beads for effective immunoprecipitation and co-immunoprecipitation.

Magnetic IP with NHS chemistry is convenient and allows for the immobilization of IP antibody from any species and subtype without the variations in coupling efficiency seen with Protein A and Protein G. The IP antibody is covalently attached so target proteins or co-IP protein complexes can be eluted and analyzed without antibody contamination. The kit contains an optimized protocol and all buffers and reagents necessary for antibody coupling and high yield IP or co-IP using either manual magnetic stands or automated platforms such as the Thermo Scientific KingFisher Instruments.

Features of the Direct Magnetic IP/Co-IP Kit:

Antibody-free IP—antibody is irreversibly attached to the magnetic beads, resulting in negligible co-elution of intact antibody or heavy and light chains with the purified antigen
Fast—complete antibody conjugation and immunoprecipitation in less than 4 hours
Convenient—the IP/co-IP kit contains magnetic beads and all essential buffers for optimal immunoprecipitation
Antibody compatible—use with any antibody species, class or isotype
Low nonspecific binding—the bead surface is pre-blocked and all nonreacted NHS-ester groups are fully quenched
Protocol-compatible—protein coupling to the beads and immunoprecipitation can be performed manually or by automation (e.g., Thermo Scientific KingFisher Instruments)
Scalable—use only the amount of antibody needed for a single IP experiment or prepare a larger quantity of antibody magnetic beads for multiple experiments

The protocol for the Pierce Direct IP/Co-IP Kit conjugates IP antibody to Pierce NHS-Activated Magnetic Beads through NHS-ester chemistry. The antibody-bound beads are then incubated with quenching buffer to block remaining active NHS sites. The prepared antibody-bound beads are incubated with cell lysate or tissue extract that contain the protein antigen of interest, allowing the antigen:antibody complex to form. The beads are washed to remove nonbound material and a low-pH elution buffer is used to dissociate bound antigen from the antibody-bound bead. Neutralization buffer is included to prevent precipitation of the isolated antigen and to ensure protein activity in downstream applications. Lane Marker Sample Buffer is included for preparing samples for SDS-PAGE analysis. The protocol is optimized for 2 to 10μg of IP antibody per assay. For optimal co-IP yields, use 10μg of antibody. The beads can be processed manually using a magnetic stand or by automation with an instrument such as the Thermo Scientific KingFisher Flex.

Traditional IP with Protein A/G Magnetic Beads, involving passive, noncovalent coupling of antibody with Protein A/G typically provides higher antigen yields than IP protocols that involve covalent attachment of antibody to magnetic beads through crosslinking or NHS-ester chemistry. Covalent attachment inevitably causes loss of some functional antibody binding sites. To overcome this limitation with NHS-chemistry, it may be necessary to use larger amounts of IP antibody in the direct IP method than in the traditional IP method. The advantage of immunoprecipitation with IP antibody covalently bound to the magnetic bead is Western blots and gels devoid of interfering antibody bands.

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Pierce™ Co-Immunoprecipitation Kit (Thermo Scientific™)

The Thermo Scientific Pierce Co-Immunoprecipitation Kit provides covalent antibody immobilization and all the components necessary to perform a properly controlled co-IP experiment without antibody interference in the final product.

Co-IP is a common approach to study protein:protein interactions that uses an antibody to immunoprecipitate the antigen (bait protein) and co-immunoprecipitate any interacting proteins (prey proteins). Traditional co-IP methods that use Protein A or G result in co-elution of the antibody heavy and light chains that may co-migrate with relevant bands, masking important results. The Pierce Co-IP Kit resolves this issue by covalently coupling antibodies onto an amine-reactive resin. The kit includes optimized buffers for protein binding and recovery, reagents to perform control experiments and efficient spin columns and collection tubes, which shorten the protocol and minimize handling and mixing.

Features of the Co-Immunoprecipitation Kit:

Works for any purified antibody—perform co-IP experiments using any purified antibody regardless of species or Ig class (chicken IgY, human IgE, mouse IgG1, IgM, etc.); no dependence on Protein A and Protein G affinity binding
No antibody interference—covalent attachment method and non-reducing elution system yields pure co-IP products without contamination by the IP antibody
Reusable antibody beads—covalent antibody immobilization allows prepared IP resin to be reused and provides potential savings of costly antibody
Complete kit—all reagents for antibody immobilization and many IP experiments, including binding and wash buffers, elution buffer and convenient microcentrifuge spin columns to make resin manipulations easier and more efficient.
Control beads included—underivatized agarose beads provided for use as negative control for nonspecific binding
Versatile—co-IP method is compatible with any physiolgical (non-denaturing) protein sample buffer that is compatible with the specific antibody-antigen and protein interaction complex.

Co-immunoprecipitation (Co-IP) is a popular in-vitro method for discovering protein interactions. The Pierce Co-Immunoprecipitation Kit is configured to provide the essential tools to effectively perform Co-IP experiments in a manner that yields pure IP and co-IP products free of troublesome IP antibody. This improvement on the traditional co-IP technique is achieved by replacing Protein A/G agarose beads with AminoLink Plus Resin to which pure IP antibodies can be directly and permanently conjugated. Gentle (non-reducing, non-denaturing) elution buffer allows dissociation of the IP targets while retaining the immobilized antibody on the agarose beads. The result is a pure product that can be analyzed by Western blotting (or even sequenced) without interference from antibody.

In addition to eliminating IP antibody contamination, the antibody-conjugation method integral to the Pierce Co-IP Kit avoids non-specific binding interactions and leaching contamination that can result from traditional Protein A and Protein G methods. Although the conjugation method requires that the starting antibody be in an amine-free buffer without BSA, gelatin or other storage protein stabilizers, the fact that it does not depend on Protein A or Protein G means that any antibody species or class can be used for IP (e.g., IgM and chicken IgY). Indeed any purified protein can be immobilized for use in affinity purification of binding partners.

Pierce™ Crosslink IP Kit (Thermo Scientific™)

The Thermo Scientific Pierce Crosslink IP Kit adapts the traditional IP method to include reagents and protocol for crosslinking IP antibodies to Protein A/G agarose to enable antigen immunoprecipitation without antibody contamination.

The primary benefits resulting from these features are the ability to purify target protein without contamination by the antibody and the ability to more effectively wash and separate samples from the beaded agarose resin.

Features of the Crosslink IP Kit:

Eliminates antibody contamination—antibody is irreversibly attached to the agarose beads so that co-elution of heavy and light chains with the purified protein is minimized
Easy and efficient scalability—use only the amount of antibody needed for a single IP experiment or immobilize 100-200 µg of antibody to prepare ready-made IP affinity resin for many experiments
Assay reliability and sample handling—Pierce Spin Columns eliminate resin loss and provide for more efficient separation of solutions than traditional IP methods that use only microcentrifuge tubes
Prepared antibody affinity resin is reusable—because the antibody is covalently immobilized and usually is not inactivated by the mild elution procedure, the resin often can be used several times
Complete kits—package includes sufficient reagents and spin columns to perform at least 50 antibody immobilizations and IP experiments

Applications:
• Immunoprecipitation followed by electrophoresis and excision of the protein for mass spectrometry or sequencing identification analysis
• Immunoprecipitation and SDS-PAGE analysis of a target protein whose molecular weight is similar to the heavy or light chain antibody fragment
• Immunoprecipitation of target proteins for subsequent analysis by nondenaturing methods (i.e., methods not involving SDS-PAGE)

The Pierce Crosslink IP Kit method involves capturing the IP antibody to Protein A/G Agarose resin and covalently immobilizing it to the support by crosslinking with disuccinmidyl suberate (DSS). The antibody resin is then incubated with the sample that contains the protein antigen of interest, allowing the antibody:antigen complex to form. After washing to remove nonbound (presumably undesired) components of the sample, the antigen is recovered by dissociation from the antibody with elution buffer supplied in the kit. The entire procedure is performed in a microcentrifuge spin cup, allowing solutions to be fully separated from the agarose resin upon brief centrifugation. Only antigen is eluted by the procedure, enabling it to be identified and further analyzed without interference from antibody fragments. Furthermore, the antibody resin often can be reused for additional rounds of immunoprecipitation.

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Pierce™ HA-Tag IP/Co-IP Kit (Thermo Scientific™)

The Thermo Scientific Pierce HA Tag IP/Co-IP Kit provides the affinity resin, positive control and other reagents necessary to perform immunoprecipitation (IP) or co-immunoprecipitation (Co-IP) reactions with an HA-tagged bait protein.

Features of the HA-Tag IP/Co-IP Kit:

Improved—updated kit includes more immobilized antibody resin and improved protocol along with GST-PI3K-SH2-HA fusion protein as a positive control
Specific—highly specific anti-HA monoclonal antibody provides high-yield immunoprecipitation products and clean Western blot detection
Compatible—includes protocols and reagents for multiple elution conditions to accommodate different protein sensitivities and downstream applications
Robust—the kit and the anti-HA agarose are compatible with various cell lysates and physiologic (non-denaturing) buffer systems
Convenient and easy—complete kit includes all necessary reagents, convenient spin columns and easy-to-follow instructions

This kit is based on crosslinked beaded agarose to which a highly specific anti-HA antibody is covalently immobilized. The ready-to-use affinity resin, together with the included buffers, microcentrifuge spin columns, positive control, and easy-to-follow instructions, constitute a complete set of reagents sufficient for 25 for IP or Co-IP assays. Upon incubation with a sample containing the tagged fusion protein, interaction complexes involving the HA-tagged bait protein are captured on the agarose beads. After simple washing steps, the specific protein interaction complex is easily eluted from the resin in the supplied elution buffer or SDS-PAGE sample loading buffer for subsequent analysis.

The hemagglutinin (HA) peptide (YPYDVPDYA), derived from the human influenza virus HA protein is one of several fusion protein tags used for recombinant protein expression. Utilizing a specific, high-affinity immobilized antibody, HA-tagged fusion proteins can be quickly purified from bacterial and mammalian cell lysates as well as from the Pierce Human in vitro translation reactions. For co-immunoprecipitation reactions, simple wash steps allow enrichment and elution of specific protein interaction complexes into the supplied elution buffer or SDS-PAGE sample loading buffer for subsequent analysis.

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Pierce™ Biotinylated Protein Interaction Pull-Down Kit (Thermo Scientific™)

The Thermo Scientific Pierce Biotinylated Protein Interaction Pull-Down Kit contains the necessary components to capture and purify interactors of a biotin-labeled protein or ligand.

Features of the Biotinylated Protein Interaction Pull-Down Kit:

• Provides a complete, affordable and easy-to-use strategy for discoverying protein:protein interactions
• Uses common laboratory equipment and reagents (e.g., microcentrifuges, mini-gels, protein stains)
• Adaptable to single- or multiple-sample demands
• Flexible pull-down format uses spin cups for easy and efficient manipulation of streptavidin agarose beads

You provide the biotinylated protein as the "bait" (see

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Pierce™ HA-Tag Magnetic IP/Co-IP Kit (Thermo Scientific™)

The Thermo Scientific Pierce Magnetic HA-Tag IP/Co-IP Kit contains specific immunoaffinity magnetic beads and reagents to perform immunoprecipitation assays of HA fusion proteins or co-IP experiments using HA-tagged bait proteins.

Features of the HA-Tag Magnetic IP/Co-IP Kit:

Specific magnetic beads—covalently immobilized high-quality anti-HA monoclonal antibody enables high yields of immunoprecipitation products
Low non-specific binding—stable, pre-blocked beads and specific antibody minimize off-target binding
Trouble-free elution—low-pH elution buffer ensures recovery of HA-tagged protein interaction complexes without antibody leaching contamination
Convenient and fast—complete kit and easy-to-follow instructions provide optimized protocols to perform IP or Co-IP experiment in approximately 1 hour
Versatile—magnetic beads are compatible with manual and automated magnetic separation workflows (e.g., Thermo Scientific KingFisher Instruments)

Unlike traditional IP procedures based on capture with Protein A/G beads, this kit uses magnetic beads containing pre-immobilized anti-HA antibody. These Pierce Anti-HA Magnetic Beads ensure specific binding of HA-tagged protein complexes from biological samples. Because the antibody is covalently attached to the beads, IP-targets are easily eluted and recovered without antibody contamination. The complete IP kit includes the magnetic beads, lysis/wash buffer, low-pH elution buffer, neutralization buffer, HA-tag positive control lysate, and non-reducing sample buffer for SDS-PAGE. Protocols are provided for both manual and automated magnetic separation workflows. Sufficient components are provided to perform 40 IP or co-IP assays.

The hemagglutinin (HA) peptide (YPYDVPDYA), derived from the human influenza virus HA protein, is one of several fusion protein tags used for recombinant protein expression. The Pierce Magnetic HA-Tag IP/Co-IP Kit uses a specific, high-affinity immobilized antibody (clone 2-2.2.14) for rapid purification of HA-tagged fusion proteins from bacterial and mammalian cell lysates, as well as from lysates prepared with the Pierce Human in vitro Translation Kits. The beads are incubated with a cell lysate containing HA-tagged protein, the fusion protein is captured, and the beads are subsequently washed and then eluted using low-pH elution buffer or non-reducing sample buffer. The protocol and buffers have been optimized for both IP and co-IP reactions, enriching for specific protein interaction complexes in the eluted samples. Anti-HA antibody can be used to detect HA-tagged protein by Western blot analysis.

For information about the binding capacity and other properties of the magnetic beads used in this kit, see Pierce Anti-HA Magnetic Beads (Part No. 88836).

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Pierce™ c-Myc-Tag Magnetic IP/Co-IP Kit (Thermo Scientific™)

The Thermo Scientific Pierce Magnetic c-Myc-Tag IP/Co-IP Kit contains specific immunoaffinity magnetic beads and reagents to perform immunoprecipitation assays of c-Myc fusion proteins or co-IP experiments using c-Myc-tagged bait proteins.

Unlike traditional IP procedures based on capture with Protein A/G beads, this kit uses magnetic beads containing pre-immobilized anti-c-Myc antibody. These Pierce Anti-c-Myc Magnetic Beads ensure specific binding of c-Myc tagged protein complexes from biological samples. Because the antibody is covalently attached to the beads, IP-targets are easily eluted and recovered without antibody contamination. The complete IP kit includes the magnetic beads, lysis/wash buffer, low-pH elution buffer, neutralization buffer, c-Myc-tag positive control lysate, and non-reducing sample buffer for SDS-PAGE. Protocols are provided for both manual and automated magnetic separation workflows. Sufficient components are provided to perform 40 IP or co-IP assays.

Features of the c-Myc-Tag Magnetic IP/Co-IP Kit:

Specific magnetic beads—covalently immobilized high-quality anti-c-Myc monoclonal antibody enables high yields of immunoprecipitation products
Low non-specific binding—stable, pre-blocked beads and specific antibody minimize off-target binding
Trouble-free elution—low-pH elution buffer ensures recovery of c-Myc-tagged protein interaction complexes without antibody leaching contamination
Convenient and fast—complete kit and easy-to-follow instructions provide optimized protocols to perform IP or Co-IP experiment in approximately 1 hour
Versatile—magnetic beads are compatible with manual and automated magnetic separation workflows (e.g., Thermo Scientific KingFisher Instruments)

The c-Myc peptide (EQKLISEEDL) derived from the C-terminus region of human c-Myc protein is one of several fusion protein tags used for recombinant protein expression. The Pierce c-Myc Magnetic IP/Co-IP Kit uses a specific, high-affinity immobilized antibody (clone 9E10) for rapid immunoprecipitation of c-Myc tagged fusion proteins from bacterial and mammalian cell lysates, as well as from lysates prepared with the Pierce Human in vitro Translation Kits. The beads are incubated with a cell lysate containing c-Myc tagged protein, the fusion protein is captured, and the beads are subsequently washed and then eluted using low-pH elution buffer or non-reducing sample buffer. The protocol and buffers have been optimized for both IP and co-IP reactions, enriching for specific protein interaction complexes in the eluted samples. Anti-c-Myc antibody can be used to detect c-Myc tagged protein by Western blot analysis.

For information about the binding capacity and other properties of the magnetic beads used in this kit, see Pierce Anti-c-Myc Magnetic Beads (Part No. 88842).

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EZ-Link™ Desthiobiotinylation and Pull-Down Kit (Thermo Scientific™)

The Thermo Scientific EZ-Link Desthiobiotinylation and Pull-Down Kit is a complete set of reagents to biotin-label purified proteins and use them to purify interaction complexes using streptavidin agarose resin with mild elution conditions.

Features of the EZ-Link Desthiobiotinylation and Pull-Down Kit:

Sulfo-NHS-LC-Desthiobiotin – label proteins with a biotin-like that specifically binds streptavidin but also easily elutes with mild conditions
Pull-down assay – simple, optimized protocol and reagents to label proteins and recover (purify) intact, functional protein interaction complexes
Complete kit – includes labeling reagent and desalting columns to prepare bait-proteins, and streptavidin agarose and elution buffer to perform pull-down

This all-in-one labeling and purification kit uses Sulfo-NHS-LC-Desthiobiotin, an amine-reactive NHS-ester reagent, to effectively tag a purified or mixed protein sample for use as bait in capturing and purifying protein interactions. The desthiobiotin tags of labeled proteins bind to streptavidin with high specificity yet readily elute with very mild conditions (i.e., by competitive displacement with regular, free biotin). The system enables effective yet gentle capture and isolation (pull-down) of specific protein interaction complexes from cell lysates using streptavidin agarose beads. The complete kit includes ready-to-use 1 mg vials of desthiobiotinylation reagent, desalting columns to process labeled protein, and high-capacity streptavidin agarose with buffers to perform several microcentrifuge-scale pull-down assay experiments with ease. Identify novel interactions between a known protein (bait) and previously undiscovered target (prey), or confirm interactions among known bait and prey proteins.

Desthiobiotin is a single-ring, sulfur-free analog of biotin that binds to streptavidin with nearly equal specificity but less affinity than biotin (1/Kd = 1011 vs. 1015M, respectively). Consequently, desthiobiotinylated bait proteins and their interacting partners can be eluted readily and specifically from streptavidin affinity resin using mild conditions based on competitive displacement with free biotin. For pull-down assay experiments with biological samples, this soft-release characteristic of desthiobiotin also helps to minimize co-purification of endogenous biotinylated molecules, which remain bound to streptavidin upon elution of the target protein complexes with free biotin. The modified avidin-biotin affinity system also eliminates the need to use harsh elution conditions that might disassociate complexes and/or damage the target protein or cell. Desthiobiotin-based techniques are ideal when using native or recombinant proteins that are not expressed with a fusion tag and when isolating captured proteins under native conditions, such as targeting intact cells or cell surface proteins.

Sulfo-NHS-LC-Desthiobiotin is a variant of biotin that is activated as a sulfo-NHS ester with a long-chain (LC) spacer arm to covalently label primary amines (-NH2) of proteins or other molecules with desthiobiotin groups. The No-Weigh Format (1 mg resealable vials) eliminates the difficulties associated with weighing small quantities of reagent and maximizes protection of unused reagent from hydrolysis.

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Pierce™ His Protein Interaction Pull-Down Kit (Thermo Scientific™)

Thermo Fisher Scientific Pierce His Tag Protein Interaction Pull-Down Kit contains the necessary components to capture and purify proteins that interact with His-tagged fusion proteins.

Features of His Tag Protein Interaction Pull-Down Kit:

6xHis pull-down (Product No. 21277)—purifies protein interactors of any His-tagged fusion protein
Complete kits—provide all components and detailed protocol for purifying protein:protein interactions
No special equipment needed—use common laboratory equipment and reagents (e.g., microcentrifuge)
Convenient—microcentrifuge spin columns facilitate simple and efficient manipulation of agarose beads, including simple processing of multiple samples
Flexible—instructions include protocols for use with bait and prey proteins expressed from a variety of sample types
Less non-specific binding—Cobalt chelate resin is more specific for histidine-tagged fusion proteins than nickel resins, resulting in less non-specific binding
Binding—Binds 10 to 25 mg of histidine-tagged fusion protein per mL of resin

Applications:

• Discover a new protein:protein interaction from a cell lysate
• Confirm a putative interaction from a cell lysate or with a previously purified protein
• Extract protein:protein interaction information from in vitro transcription/translation lysates

You provide the tagged fusion protein as the "bait" and the cells expressing the putative protein interaction target ("prey"), and the Pull-Down Kits provide everything else: cell lysis buffer, microcentrifuge spin columns, tag-specific affinity resin (agarose beads) and optimized buffers and protocol. The Pull-Down Kits are designed to teach the method to first-time users and to increase ease-of-use, convenience and reproducibility for experienced researchers.

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Pierce™ Direct IP Kit (Thermo Scientific™)

The Thermo Scientific Pierce Direct IP Kit uses an activated resin to covalently immobilize IP antibodies (any species or class) on agarose beads without the aid of Protein A/G, enabling immunoprecipitation without antibody interference.

The primary benefits resulting from this method are the opportunity to use any species or subclass of purified antibody (not just types that bind to Protein A or G) and the ability to purify target protein without contamination by the antibody. The method also makes it possible to immunoprecipitate antigens from serum samples without co-purifying non-target immunoglobulins. Finally, the kit uses microcentrifuge spin cups to effectively wash and separate samples from the beaded agarose resin.

Features of Direct IP Kit:

Requires less than 10 µg of antibody—optimized procedure requires only 2 to 10 µg of antibody for a single IP experiment, no more than is required for traditional IP methods
Easy and efficient scalability—use only the amount of antibody needed for a single immunoprecipitation or immobilize 100 to 200 µg of antibody to prepare ready-made IP affinity resin for many experiments
Minimal antibody contamination—antibody is irreversibly attached to the agarose beads so that co-elution of heavy and light chains with the purified protein is minimized (i.e., less than 5%)
IP with any species and subclass of antibody—Use chicken IgY, human IgE, mouse IgM or any other purified protein
Prepared antibody affinity resin is reusable—because the antibody is covalently immobilized and not inactivated by the mild elution procedure, the resin often can be used several times
Convenient sample handling—spin columns (see dimensions) eliminate resin loss and provide for efficient separation of solutions
Complete kit—package includes cell lysis buffer and sufficient reagents and spin columns to perform at least 50 antibody immobilizations and IP experiments
Requires purified antibody—IP antibody solution must be free of BSA, gelatin or other stabilizer proteins; for easy removal of these proteins, see our Antibody Clean-up Kit (Part No. 44600)

Applications:
• Immunoprecipitation followed by electrophoresis and excision of the protein for mass spectrometry or sequencing identification analysis
• Immunoprecipitation and SDS-PAGE analysis of a target protein whose molecular weight is similar to the heavy or light chain antibody fragment
• Immunoprecipitation of target proteins for subsequent analysis by nondenaturing methods (i.e., methods not involving SDS-PAGE)
• Immunoprecipitation with antibodies that do not bind to Protein A, Protein G or Protein A/G
• Immunoprecipitation from serum and other sample that contain immunoglobulins

Direct immunoprecipitation in two easy steps:

Step 1: Attach antibody to agarose resin: The first step in the Direct IP Method is to prepare the IP antibody affinity resin. Pure antibody (see Highlights) is incubated in phosphate buffer with an appropriate volume of AminoLink Plus Coupling Resin, an aldehyde-activated beaded agarose resin. During the incubation, various lysine epsilon-amines on the surface of the large antibody molecule react with aldehyde groups on the resin to form Schiff-base bonds that are then stabilized to secondary amine bonds in the presence of cyanoborohydride. The immobilization reaction, known as reductive amination, is nondenaturing and typically results in greater than 85% antibody coupling efficiency with minimal loss of antigen-binding function.

Step 2: Perform immunoprecipitation experiments: Once prepared, the antibody resin can be used in several parallel aliquots or in total for IP experiments. Antibody resin is incubated with a sample that contains the protein antigen of interest, allowing the antibody:antigen complex to form. After washing to remove nonbound (presumably undesired) components of the sample, the antigen is recovered by dissociation from the antibody with elution buffer supplied in the kit. The entire procedure is performed in a microcentrifuge spin cup, allowing solutions to be fully separated from the agarose resin upon brief centrifugation. Only antigen is eluted by the procedure, enabling it to be identified and further analyzed without interference from antibody fragments. Furthermore, the antibody resin can be reused for additional rounds of immunoprecipitation.

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Pierce™ Classic IP Kit (Thermo Scientific™)

The Thermo Scientific Pierce Classic IP Kit uses high-quality Protein A/G agarose beads, microcentrifuge spin columns and buffers to perform traditional immunoprecipitation assays with ease.

The kit uses high-capacity Protein A/G agarose affinity resin for efficient binding of most species and subclasses of IP antibodies. The included IgG elution buffer provides milder and less denaturing recovery of antibody:antigen complexes than the traditional method of boiling in reducing sample buffer for SDS-PAGE, facilitating a greater variety of methods for subsequent analysis. The microcentrifuge spin column format helps to ensure effective washing and separation of samples from the beaded agarose affinity resin.

Features of the Classic IP Kit:

Assay consistency—spin columns (see dimensions) eliminate resin loss and provide for more efficient separation of solutions than traditional IP methods that use only microcentrifuge tubes
Fast—immunoprecipitate (IP) in less than one hour
Mild elution conditions—recover antigen without harsh detergents or reducing agents
Easy-to-follow instructions—purify target protein from crude lysate in four simple steps
Suitable for most common antibody types—Protein A/G provides excellent binding for most mouse, rabbit, human and goat IgG subclasses
Complete kit—includes lysis/wash buffer, binding and elution buffers, control agarose resin, sample loading buffer and spin columns

Applications:
• Routine, traditional immunoprecipitation experiments
• Immunoprecipitation for analysis in nonreducing conditions

Like traditional IP methods, the Pierce Classic IP Kit procedure involves formation of antibody:antigen complexes in a sample solution and then capture of that complex to an IgG-binding protein that is covalently bound to beaded agarose resin (Protein A/G Agarose). After washing to remove nonbound (presumably undesired) components of the sample, the antigen and antibody are recovered from the beaded resin with elution buffer supplied in the kit. The entire procedure is performed in a microcentrifuge spin column, allowing solutions to be fully separated from the agarose resin upon brief centrifugation.

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Pierce™ GST Protein Interaction Pull-Down Kit (Thermo Scientific™)

Thermo Fisher Scientific Pierce GST Tag Protein Interaction Pull-Down Kits contains the necessary components to capture and purify proteins that interact with GST-tagged fusion proteins.

Features of GST Tag Protein Interaction Pull-Down Kit:

GST pull-down (Product No. 21516)—gently purifies protein interactors of any GST-tagged fusion protein without denaturing
Complete kits—provide all components and detailed protocol for purifying protein:protein interactions
No special equipment needed—use common laboratory equipment and reagents (e.g., microcentrifuge)
Convenient—microcentrifuge spin columns facilitate simple and efficient manipulation of agarose beads, including simple processing of multiple samples
Flexible—instructions include protocols for use with bait and prey proteins from many different sources
Binding—Binds approx. 8 mg of GST-tagged fusion protein per mL of resin

Applications:
• Discover a new protein:protein interaction from a cell lysate
• Confirm a putative interaction from a cell lysate or with a previously purified protein
• Extract protein:protein interaction information from in vitro transcription/translation lysates

You provide the tagged fusion protein as the "bait" and the cells expressing the putative protein interaction target ("prey"), and the Pull-Down Kits provide everything else: cell lysis buffer, microcentrifuge spin columns, tag-specific affinity resin (agarose beads) and optimized buffers and protocol. The Pull-Down Kits are designed to teach the method to first-time users and to increase ease-of-use, convenience and reproducibility for experienced researchers.

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InsuQuant Mass Spectrometric Kit (Thermo Scientific™)

Simplify your affinity purification workflow with the Thermo Scientific™ InsuQuant™ Mass Spectrometric Kit, an exclusive pre-analytical solution designed to simultaneously detect, differentiate, and quantify endogenous and exogenous insulin types.