First – what is protein labeling?
Protein labeling involves the utilization of appropriate molecular labels in order to purify or detect the labeled proteins, along with its binding partners. Proteins are known to be one of the largest and critical biomolecules present in living organisms, since they perform vital functions. Once formed, proteins within a cell carry out interactions with various other biomolecules so that the cell can be kept alive. As a result, it has become very important that specialized areas of study such as protein chemistry and proteomics are undertaken extensively for researching the mechanism of protein binding.
A number of strategies can be implemented in order to carry out successful protein labeling, with these strategies leading to covalent attachment of molecules such as reporter enzymes, biotin, radioactive isotopes, and fluorophores, to the target protein or nucleotide sequence. The demand for disease diagnosis, prevention, and treatment has been ever-increasing across the globe, in line with the growing incidences of chronic diseases such as cancer, which has created a sustained demand for protein labeling techniques. Additionally, in the area of proteomic research, there has been an increase in spending on research and development activities in recent years that has aided the progress of protein analysis, thus driving the demand for these techniques.
Protein labeling – where it is used?
Protein labeling finds application in a number of areas, which include fluorescence assays, immunological techniques, mass spectrometry, cell-based assays, and protein microarray. Immunological techniques such as immunoassays, flow cytometry, immunofluorescence, and western blotting generally make use of labels. Besides being used in research for cancer and autoimmune disease, immunological techniques are also used to detect diabetes, rheumatoid arthritis, neurological and gastrointestinal diseases, systemic lupus erythematosus, and pulmonary diseases. On the other hand, the demand for fluorescence microscopy has also witnessed strong growth in recent years, primarily due to the substantial development of high-throughput fluorescence microscopy for an in-depth biological analysis in academic research of cellular pathways.
There are both in-vitro and in-vivo protein labeling methods involved in the industry, with the former being more widely utilized across the globe. The in-vitro methods include enzymatic labeling, co-translational labeling, nanoparticle labeling, dye-based labeling, and site-specific labeling, among others. On the other hand, in-vivo methods include radioactive labeling and photoreactive labeling. In the in-vitro segment, enzymatic labeling generates the majority of the revenue for the industry, while nanoparticle labeling usage is expected to become very widespread in the coming years.
Looking into the types of in-vitro protein labeling methods
Enzymatic protein labeling is a site-specific modification method which is robust and highly efficient under mild reaction conditions, with a number of major developments having been seen in this area over the past decade. Enzymes are notable for being efficient catalysts in chemical reactions, and features such as rapid reaction rates, high specificity, and the ability to work even under mild reaction conditions have made them a noteworthy option in the protein labeling industry. The major classes of enzymes used include transferases, peptidases, oxidoreductases, and ligases; other classes used on a more limited scale include tyrosinase, peroxidase, and glycan synthesis-involved enzymes. Site-specific protein labeling and engineering can be achieved due to the incorporation of bio-orthogonal functionalities into proteins, brought about by the covalent site-specific enzyme binding.
The use of fluorescent dyes has become widespread in recent years in areas such as immunofluorescence, flow cytometry, and Western blotting. These dyes are synthetic or natural compounds, which absorb electromagnetic or light energy, and then re-emit it at a lower energy, in photonic form. Fluorescent dyes conventionally belong to either of the following 3 groups – fluorescent dyes, fluorescent proteins, and quantum dots.
Additionally, various fluorescent dyes are being developed and optimized for specific instrumentations and applications, in conjunction with the evolution of various advanced fluorescence technologies in recent years. The use of fluorescent dyes in protein labeling offers advantages such as high sensitivity even for small sample sizes; wide linear dynamic range; significantly lower interference than spectrophotometric measurements as very few materials have fluorescing ability; and high throughput.
The utilization of site-specific protein labeling using a number of probes at internal sites and the generation of defined protein-protein conjugates is expected to greatly aid the study of biological function of proteins, as well as in the development of diagnostic and therapeutic bioconjugates. In the past few years, there has been a constant focus on developing strategies in order to achieve selective protein labeling in living cells. For instance, the incorporation of unnatural amino acids in site-specific protein labeling is being looked at with great interest, as it aids in attaching various organic probes to a specific protein position more precisely. The use of genetically encoded tags for individual proteins, chemical labeling by using N-hydroxysuccinimidyl (NHS) esters and maleimides. Site-specific covalent protein labeling using fluorophores as well as other moieties has enabled the development of a variety of assays for the purpose of proteome analysis.
The use of nanoparticle-based in-vitro protein labeling method is expected to gather rapid pace in the coming years. The method involves the use of nanoparticles, with gold nanoparticles being particularly utilized for linking specifically to hinge-thiol or antibody on Immunoglobin G (IgG) or fragment antigen-binding (Fab). Nanoparticles, on account of their small size, can reach intricate locations and produce covalent bonds with the target protein or peptide; this enables better stability and labeling. The use of nanoparticles in labeling offers high stability, besides preventing interactions between metal and antibody, and providing high sensitivity during protein quantification and detection. Some of the most common strategies when using gold nanoparticles for peptide molecule labeling include covalent conjugation and classic passive adsorption. An advantage of this method is that it becomes easy to carry out R&D and manufacturing process of diagnostic kits, owing to the convenient conjugation of gold nanoparticles.
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Protein labeling is in high demand, particularly in Asia Pacific
The demand for protein labeling has been high in regions such as North America, where there has been a significant growth in the number of research processes that have employed fluorescence microscopy, ELISA, immunological techniques, and Western blotting. The growth in the number of disease diagnosis-related studies, and the consequent adoption of protein labeling techniques, is expected to additionally promote the regional market development. The Asia Pacific region is expected to witness substantial growth in the coming years on account of the vast number of ongoing proteomics-related research projects in the region.
Lastly – some notable developments in this industry
There are a number of companies involved in the development of protein labeling products and techniques, including Thermo Fisher Scientific, Merck, New England Biolabs, GE Healthcare, Promega Corp., Perkin Elmer, LGC, and Jena Bioscience, among others. Besides these organizations, a number of medical and research institutes are also investing their efforts in driving developments in this industry. In February 2022, EpiCypher announced a partnership with New England Biolabs regarding the co-development of the CUT&RUN Library Prep Kit. The product is being developed for optimal performance with EpiCypher’s CUTANA CUT&RUN epigenomic assays, with the product being powered by NEBNext reagents from New England Biolabs.
Besides product developments and technological advancements, other strategies such as mergers and acquisitions, geographical expansions, and partnerships with research institutes are also being explored by competitors in this space. For instance, in December 2021, Battery Ventures announced the acquisition of LI-COR Biosciences, a notable manufacturer of instrumentation systems in the areas of environmental research, drug discovery, protein research, and therapeutics development.
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