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Reagents for Deciphering the Interactome

Exploring PPI applications

The human interactome has been estimated to cover approximately 400,000 protein-protein interactions (PPI). Understanding and modulating these biomolecular complexes is one of the foremost challenges in the discovery of innovative drugs for pathologies related to oncology, inflammation, CNS, metabolism, and infectious diseases, among others.

Many possibilities exist, including Receptor/ligand, Nuclear receptor/ligand, Protein/DNA, Protein/RNA, Protein/peptide, Protein/carbohydrates, and many more.

Learn how researchers use Revvity’s immunoassays to build various PPI assays.

For research use only. Not for use in diagnostic procedures.

Reagents for Deciphering the Interactome

Virus blockade assay

protein-protein-interaction-lookbook-ppi-virus-blockade-assay

Adapted from Newton et al. / Journal of Biomolecular Screening 18(3) (2012) 237–246.


A wide range of biomolecular interactions can be assessed

The advances in identifying short primary sequences mostly involved in the PPI interfaces, also called hot-spots, have greatly facilitated the setup of biochemical assays to identify PPI modulators. However in some projects the binding epitopes involved in PPI interfaces are not fully understood, which requires the use of full size proteins to reproduce the interaction and screen for inhibitors or stabilizers.

HTRF is a popular technology for PPIs, whatever the molecular weight of your protein partners, as illustrated below: 

protein-protein-interaction-lookbook-ppi-large-size-complex_ppi-large-size-complex

Large size complex
B-Catenin/TCF4

This study was performed to show the ability of HTRF PPI reagents to address large biomolecular interactions. Recombinant human GST-β-Catenin (115 KDa) and 6HIS-TCF4 (55 KDa) interaction was detected using Anti GST-Eu Cryptate (150 KDa) and Anti 6HIS-d2 (150 KDa) antibodies.

HTRF is suitable for low to high affinity interactions

Protein-Protein interaction networks are extended and complex. They are also conveyed by a vast number of non-covalent binding modes. This diversity results in a wide range of affinities from very weak and transient interactions to very strong (High µM to picomolar).

The reliability of HTRF, coupled with a wide diversity of PPI tools, helps to enable the efficient measurement of complex interactions displaying low to high Kd.

HTRF PPI assays taken from published papers, displaying a large panel of affinities. 

protein-protein-interaction-lookbook-ppi_ppi-large-panel-of-affinities

Let’s see how to get started with your own PPI!
  1. Choose your target interaction

    Receptor/ligands, RNA, DNA, lipids, enzymes, etc. Any interacting proteins are within your reach: It’s all about what you want!

  2. Select assay format & starting material

    Antibodies, native partner, biotinylated partner, tagged protein, Fc fused proteins matching your goals.

  3. Choose your detection reagents

    46 options ensure that you find a match for your interaction: labeling kits, affinity reagents, detection and anti species reagents.

  4. Assay optimization and data processing

    A single optimization step to adjust reagents to each other.
    And off to the reader you go!

Receptor-Ligand binding assay

protein-protein-interaction-lookbook-ppi-receptor-ligand-binding-assay

Adapted from C J. Rossant / Journal of Biomolecular Screening 2015, Vol. 20(4) 508–518
 


A wide range of biomolecular interactions can be assessed

The advances in identifying short primary sequences mostly involved in the PPI interfaces, also called hot-spots, have greatly facilitated the setup of biochemical assays to identify PPI modulators. However in some projects the binding epitopes involved in PPI interfaces are not fully understood, which requires the use of full size proteins to reproduce the interaction and screen for inhibitors or stabilizers.

HTRF is a popular technology for PPIs, whatever the molecular weight of your protein partners, as illustrated below: 

protein-protein-interaction-lookbook-ppi-large-size-complex_ppi-large-size-complex

Large size complex
B-Catenin/TCF4

This study was performed to show the ability of HTRF PPI reagents to address large biomolecular interactions. Recombinant human GST-β-Catenin (115 KDa) and 6HIS-TCF4 (55 KDa) interaction was detected using Anti GST-Eu Cryptate (150 KDa) and Anti 6HIS-d2 (150 KDa) antibodies.

HTRF is suitable for low to high affinity interactions

Protein-Protein interaction networks are extended and complex. They are also conveyed by a vast number of non-covalent binding modes. This diversity results in a wide range of affinities from very weak and transient interactions to very strong (High µM to picomolar).

The reliability of HTRF, coupled with a wide diversity of PPI tools, helps to enable the efficient measurement of complex interactions displaying low to high Kd.

HTRF PPI assays taken from published papers, displaying a large panel of affinities. 

protein-protein-interaction-lookbook-ppi_ppi-large-panel-of-affinities

Let’s see how to get started with your own PPI!
  1. Choose your target interaction

    Receptor/ligands, RNA, DNA, lipids, enzymes, etc. Any interacting proteins are within your reach: It’s all about what you want!

  2. Select assay format & starting material

    Antibodies, native partner, biotinylated partner, tagged protein, Fc fused proteins matching your goals.

  3. Choose your detection reagents

    46 options ensure that you find a match for your interaction: labeling kits, affinity reagents, detection and anti species reagents.

  4. Assay optimization and data processing

    A single optimization step to adjust reagents to each other.
    And off to the reader you go!

mRNA/Protein binding assay

protein-protein-interaction-lookbook-ppi-mRNA-protein-binding-assay

Adapted from M.D.Disney et al. / ACS Chem Biol. 2012 October 19; 7(10): 1711–1718 
 


A wide range of biomolecular interactions can be assessed

The advances in identifying short primary sequences mostly involved in the PPI interfaces, also called hot-spots, have greatly facilitated the setup of biochemical assays to identify PPI modulators. However in some projects the binding epitopes involved in PPI interfaces are not fully understood, which requires the use of full size proteins to reproduce the interaction and screen for inhibitors or stabilizers.

HTRF is a popular technology for PPIs, whatever the molecular weight of your protein partners, as illustrated below: 

protein-protein-interaction-lookbook-ppi-large-size-complex_ppi-large-size-complex

Large size complex
B-Catenin/TCF4

This study was performed to show the ability of HTRF PPI reagents to address large biomolecular interactions. Recombinant human GST-β-Catenin (115 KDa) and 6HIS-TCF4 (55 KDa) interaction was detected using Anti GST-Eu Cryptate (150 KDa) and Anti 6HIS-d2 (150 KDa) antibodies.

HTRF is suitable for low to high affinity interactions

Protein-Protein interaction networks are extended and complex. They are also conveyed by a vast number of non-covalent binding modes. This diversity results in a wide range of affinities from very weak and transient interactions to very strong (High µM to picomolar).

The reliability of HTRF, coupled with a wide diversity of PPI tools, helps to enable the efficient measurement of complex interactions displaying low to high Kd.

HTRF PPI assays taken from published papers, displaying a large panel of affinities. 

protein-protein-interaction-lookbook-ppi_ppi-large-panel-of-affinities

Let’s see how to get started with your own PPI!
  1. Choose your target interaction

    Receptor/ligands, RNA, DNA, lipids, enzymes, etc. Any interacting proteins are within your reach: It’s all about what you want!

  2. Select assay format & starting material

    Antibodies, native partner, biotinylated partner, tagged protein, Fc fused proteins matching your goals.

  3. Choose your detection reagents

    46 options ensure that you find a match for your interaction: labeling kits, affinity reagents, detection and anti species reagents.

  4. Assay optimization and data processing

    A single optimization step to adjust reagents to each other.
    And off to the reader you go!

Antibody epitope mapping

protein-protein-interaction-lookbook-ppi-antibody-epitope-mapping

Adapted from C J. Rossant / Journal of Biomolecular Screening 2015, Vol. 20(4) 508–518 
 


A wide range of biomolecular interactions can be assessed

The advances in identifying short primary sequences mostly involved in the PPI interfaces, also called hot-spots, have greatly facilitated the setup of biochemical assays to identify PPI modulators. However in some projects the binding epitopes involved in PPI interfaces are not fully understood, which requires the use of full size proteins to reproduce the interaction and screen for inhibitors or stabilizers.

HTRF is a popular technology for PPIs, whatever the molecular weight of your protein partners, as illustrated below: 

protein-protein-interaction-lookbook-ppi-large-size-complex_ppi-large-size-complex

Large size complex
B-Catenin/TCF4

This study was performed to show the ability of HTRF PPI reagents to address large biomolecular interactions. Recombinant human GST-β-Catenin (115 KDa) and 6HIS-TCF4 (55 KDa) interaction was detected using Anti GST-Eu Cryptate (150 KDa) and Anti 6HIS-d2 (150 KDa) antibodies.

HTRF is suitable for low to high affinity interactions

Protein-Protein interaction networks are extended and complex. They are also conveyed by a vast number of non-covalent binding modes. This diversity results in a wide range of affinities from very weak and transient interactions to very strong (High µM to picomolar).

The reliability of HTRF, coupled with a wide diversity of PPI tools, helps to enable the efficient measurement of complex interactions displaying low to high Kd.

HTRF PPI assays taken from published papers, displaying a large panel of affinities. 

protein-protein-interaction-lookbook-ppi_ppi-large-panel-of-affinities

Let’s see how to get started with your own PPI!
  1. Choose your target interaction

    Receptor/ligands, RNA, DNA, lipids, enzymes, etc. Any interacting proteins are within your reach: It’s all about what you want!

  2. Select assay format & starting material

    Antibodies, native partner, biotinylated partner, tagged protein, Fc fused proteins matching your goals.

  3. Choose your detection reagents

    46 options ensure that you find a match for your interaction: labeling kits, affinity reagents, detection and anti species reagents.

  4. Assay optimization and data processing

    A single optimization step to adjust reagents to each other.
    And off to the reader you go!

Protease assay

protein-protein-interaction-lookbook-ppi-protease-assay

Ingo H. Engels et al. / Analytical Biochemistry 390 (2009) 85–87 
 


A wide range of biomolecular interactions can be assessed

The advances in identifying short primary sequences mostly involved in the PPI interfaces, also called hot-spots, have greatly facilitated the setup of biochemical assays to identify PPI modulators. However in some projects the binding epitopes involved in PPI interfaces are not fully understood, which requires the use of full size proteins to reproduce the interaction and screen for inhibitors or stabilizers.

HTRF is a popular technology for PPIs, whatever the molecular weight of your protein partners, as illustrated below: 

protein-protein-interaction-lookbook-ppi-large-size-complex_ppi-large-size-complex

Large size complex
B-Catenin/TCF4

This study was performed to show the ability of HTRF PPI reagents to address large biomolecular interactions. Recombinant human GST-β-Catenin (115 KDa) and 6HIS-TCF4 (55 KDa) interaction was detected using Anti GST-Eu Cryptate (150 KDa) and Anti 6HIS-d2 (150 KDa) antibodies.

HTRF is suitable for low to high affinity interactions

Protein-Protein interaction networks are extended and complex. They are also conveyed by a vast number of non-covalent binding modes. This diversity results in a wide range of affinities from very weak and transient interactions to very strong (High µM to picomolar).

The reliability of HTRF, coupled with a wide diversity of PPI tools, helps to enable the efficient measurement of complex interactions displaying low to high Kd.

HTRF PPI assays taken from published papers, displaying a large panel of affinities. 

protein-protein-interaction-lookbook-ppi_ppi-large-panel-of-affinities

Let’s see how to get started with your own PPI!
  1. Choose your target interaction

    Receptor/ligands, RNA, DNA, lipids, enzymes, etc. Any interacting proteins are within your reach: It’s all about what you want!

  2. Select assay format & starting material

    Antibodies, native partner, biotinylated partner, tagged protein, Fc fused proteins matching your goals.

  3. Choose your detection reagents

    46 options ensure that you find a match for your interaction: labeling kits, affinity reagents, detection and anti species reagents.

  4. Assay optimization and data processing

    A single optimization step to adjust reagents to each other.
    And off to the reader you go!

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