Protein Interaction Analysis

Protein-protein interactions refer to the process where two or more protein molecules bind together to form a protein complex. Generally, proteins rarely function alone; they usually work in coordination with other protein molecules to achieve complex cellular functions. Therefore, focusing on the study of protein interactions is more conducive to exploring the occurrence and development of diseases, finding specific disease prediction and treatment methods, and providing new ideas for new drug development research. There are many methods for protein interaction analysis, and BiotechPack offers protein interaction analysis services, as well as subsequent mass spectrometry identification and analysis services for protein/protein mixtures in purified samples such as IP, Co-IP samples, and GST fusion protein pull-down based on LC-MS/MS technology. For more details, see:
Protein Interaction Mass Spectrometry Analysis
Protein Interaction Analysis using Co-Immunoprecipitation (Co-IP) combined with Mass Spectrometry
Protein Interaction Analysis using GST Fusion Protein Pull-down combined with Mass Spectrometry
Protein Interaction Analysis using SILAC combined with Co-Immunoprecipitation and Mass Spectrometry

1. Immunoprecipitation (Co-IP) Protein Interaction Analysis

Immunoprecipitation (Co-IP) is a method that uses the specificity between antigens and antibodies to detect physiological interactions between proteins. To maximally preserve protein interactions within cells during experiments, non-ionic detergents are used to lyse cells. When cells are lysed, intracellular protein interactions are preserved. Specific antibodies are added to bind to specific proteins in the cells, then protein A/G is added to form an antibody-protein complex. This complex is centrifuged and separated to precipitate the antibody-protein complex, discarding the supernatant. Finally, mass spectrometry is used to identify the precipitated proteins, proving the interaction between them. This method is only suitable for analyzing stable or strong protein interactions and is usually used to identify interactions between a target protein and another protein or to find unknown proteins interacting with a target protein.

BiotechPack Biotechnology offers optimized solutions for sample protein extraction, expression, and purification to ensure high-quality bait and prey proteins, providing customers with quality Co-IP protein interaction analysis services and protein mass spectrometry analysis of proteins obtained through immunoprecipitation (Co-IP).

 

BiotechPack Protein Interaction Analysis

• Advantage: Can isolate protein complexes interacting in their natural state;
• Disadvantage: Low sensitivity; cannot analyze and detect low-affinity and transient protein interactions.

2. GST Fusion Protein Pull-down Technique for Protein Interaction Analysis

The GST fusion protein pull-down technique involves inserting a vector with a GST tag into the target protein A through genetic recombination, allowing glutathione-coated beads to bind with the target fusion protein. After adding cell lysate, interacting proteins are adsorbed by the fusion protein. Excess glutathione is then added for elution, followed by MS analysis. This method is mainly used to study strong or stable protein interactions in vitro, identify possible direct interactions between two known proteins of interest, and find unknown proteins that may interact with the target protein.

As a technology-based scientific research service company, BiotechPack Biotechnology has years of experience in protein interaction analysis services and protein mass spectrometry identification analysis based on mass spectrometry. BiotechPack offers GST pull-down protein interaction analysis services and pull-down target protein mass spectrometry identification services. We feature strict quality control and short detection cycles, providing customizable one-stop services for clients, covering every step from gene synthesis to data reporting in your project.
• Advantage: Can eliminate interference from other cellular conditions; allows direct detection of protein-protein interactions; simple method, easy to operate.
• Disadvantage: Conducted outside of cells, it cannot accurately reflect intracellular protein interaction relationships.

3. SILAC with Co-Immunoprecipitation and Mass Spectrometry for Protein Interaction Analysis

Using the SILAC method, label experimental and control group cells, then perform Co-IP experiments. This relies on the specificity of antigen-antibody reactions to isolate immune complexes. LC-MS/MS is then used for qualitative and quantitative detection of proteins in the immune complexes. If a protein shows statistically significant differences between experimental and control groups, it is determined to interact with the protein under study, greatly reducing the possibility of false positive results in protein interactions. SILAC and Co-IP-MS combination can be used for high-throughput quantitative analysis of protein interaction networks under specific conditions.

BiotechPack Biotechnology uses Thermo Fisher's Q ExactiveHF mass spectrometry platform, Orbitrap Fusion mass spectrometry platform, and Orbitrap Fusion Lumos mass spectrometry platform combined with Nano-LC to provide SILAC with Co-IP and Mass Spectrometry interaction analysis service packages (SILAC-based CoIP-MS). You only need to specify your experimental goals and send us your cells, and we will handle all subsequent aspects of the project, including cell culture, cell labeling, protein extraction, antibody IP, efficiency detection, protein digestion, peptide separation, mass spectrometry analysis, raw mass spectrometry data analysis, and bioinformatics analysis.

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