Ouqiao IPAC2Protein Aggregation Counting Analyzer

How is protein composed?

Proteins are biomolecules composed of amino acids as the basic unit. The sequence of amino acids on protein molecules and the resulting three-dimensional structure constitute the diversity of protein structures. Proteins have primary, secondary, tertiary, and quaternary structures, and the structure of protein molecules determines their function.

Primary structure: The arrangement order of amino acids in protein polypeptide chains, as well as the position of disulfide bonds.

Secondary structure: The way in which a polypeptide chain wraps and folds in a certain direction within the local region of a protein molecule.

Tertiary structure: The spatial conformation of a protein's secondary structure, which is curled and folded into a specific spherical molecular structure through various secondary bonds.

Fourth order structure: The three-dimensional structure of a protein formed by the appropriate aggregation of peptide chains with tertiary structures in a multi subunit protein molecule.

The amino acid sequence of proteins is encoded by corresponding genes. In addition to the 20 "standard" amino acids encoded by the genetic code, certain amino acid residues in proteins can also be post-translational modified to undergo chemical structural changes, thereby activating or regulating proteins. However, if non-specific binding occurs between proteins, not only does the protein lose its expected activity, but it is also prone to forming inclusion bodies, leading to an increase in the cost of protein genetic engineering. The structure of aggregates includes amyloid fiber structure and inclusion body structure.

Why is it necessary to study and observe protein aggregates in biomedical engineering?

Protein aggregation has become a research hotspot in the fields of medicine and biology, where proteins exist in non natural conformations and are often accompanied by an increase in beta folding. Both Alzheimer's disease and type II diabetes are associated with protein aggregation. Studying protein aggregates helps to understand the formation of molecular diseases in vivo.

In the post genomic era of proteins and drugs, finding optimized conditions for protein crystals has always been the goal of crystal growth workers. The changes in the state of protein aggregates formed in the pre nucleation solution, including their size, morphology, and even conformation, directly affect the nucleation process. Therefore, studying the changes in the state of disordered aggregates can help analyze the conditions for the appearance of ordered aggregates and provide suitable conditions for protein crystal growth.

The State Key Laboratory of Biomembrane and Membrane Bioengineering, School of Life Sciences, Tsinghua University has purchased the "Multi angle Laser Light Scattering gel Chromatographic System" for the morphological analysis of biological sample solution, the determination of molecular weight and distribution of proteins and their aggregates, the analysis of protein homogeneity and stability, and the screening of their crystalline states and conditions, to study the impact of protein aggregates on the membrane pollution process.

Protein aggregation seriously affects drugs developed based on proteins. In pharmaceutical formulations, protein aggregation affects drug efficacy in terms of biological activity and immunogenicity. Protein aggregation occurs at various stages of the production process, including cell culture, purification, production, storage, and transportation. The pharmaceutical industry hopes to find new methods in biotechnology that can be used to detect, track, and quantitatively analyze factors that affect protein aggregation. In recent years, protein aggregates that exist in a freeze-dried stable form have been used as standard samples for quantitative detection of protein aggregates, coupled with the novel ProteoStat that can be tested in an enzyme-linked immunosorbent assay (ELISA) reader ® protein aggregation assay， Protein detection methods can be optimized.

Scientists are currently unsure why incorrect protein forms and clustering are important markers of neurodegenerative diseases, including amyotrophic lateral sclerosis (ALS), Alzheimer's disease, and mad cow disease. In a research report published in the November 1st issue of Molecular Cell, researchers from Yale University revealed the formation process of aggregates of incorrect forms of proteins by studying the pathogenesis of diseases in bacteria. Proteins are encoded and controlled by DNA and formed in cells through the assembly of ribosomes. However, sometimes proteins are not properly assembled, and these misfolded proteins tend to aggregate. The aggregation phenomenon of misfolded proteins is particularly evident in the brains of Alzheimer's patients. A research team from Yale has revealed that the antibiotic streptomycin can induce protein aggregation in Escherichia coli. Using large-scale proteomics and genetic screening techniques, researchers analyzed protein aggregation phenomena and screened bacterial proteins that can make Escherichia coli resistant to antibiotics. Eventually, researchers discovered how a special protein in bacteria protects bacteria from the pressure of hydrogen peroxide and how this protein weakens protein aggregation caused by streptomycin stimulation.

Direct visualization, size measurement, and counting of aggregates in proteins: Detecting protein aggregation status is crucial for understanding the stability and efficacy of biopharmaceutical products. When there are protein aggregates, they have a significant impact on the quality, biological activity, and immunogenicity of the product. Many aggregates will be arranged according to size and characteristics when encountering biological samples (such as soluble and insoluble, covalent and non covalent, or reversible and irreversible). The range of protein aggregates is wide, ranging from small oligomers (nanoscale) to insoluble micrometer sized aggregates composed of millions of monomer units.

Protein aggregates may be generated at any step of the manufacturing process (cell culture, purification, formation), storage, distribution, and product processing. It may be caused by various pressures, such as stirring, exposure to pH, temperature, ionic strength, or various interfaces (such as gas-liquid interfaces). In the case of high protein content (such as the formation of monoclonal antibodies), there is a possibility of further increase in aggregates. Therefore, in the development

Careful description and control of aggregates are necessary during manufacturing, subsequent storage of drugs, and product description. Similarly, it can be achieved by monitoring the status of aggregates and modifying or optimizing the production process.

Now FC-200S-IPAC provides a blue light based image based submicron particle tracking and analysis system, which can directly observe and count individual nanoscale particles (such as protein aggregates) in real-time in the liquid phase, and obtain high-resolution particle size distribution maps. This technology has the characteristics of fast, reliable, and much lower cost than scanning electron microscopy, which makes it a good alternative or supplement to existing nanoparticle analysis methods such as DLS (dynamic light scattering, also known as photon correlation spectroscopy PCS) or electron microscopy (EM)