Protein Analysis FAQ

• • Why Is a Support Medium Necessary for Protein Electrophoresis

  Protein electrophoresis relies on support media, such as polyacrylamide gels, which play essential roles in facilitating protein separation and analysis, preserving protein stability, enabling visualization, regulating electrophoresis velocity, and safeguarding samples. The support medium contributes to several critical aspects during electrophoresis: 1. Separation of Proteins The support medium provides a matrix that enables proteins to be separated based on their size, shape, or charge. Under the ........

• • How to Perform Western Blot for Ultra-High Molecular Weight Proteins (330 kDa)

  When conducting Western Blot analysis for ultra-high molecular weight proteins (e.g., 330 kDa), it is often necessary to optimize experimental conditions to improve protein separation and transfer efficiency: 1. Sample Preparation Since ultra-high molecular weight proteins migrate more slowly through the gel, it is recommended to employ a higher concentration of reducing agents (such as DTT or β-mercaptoethanol) during sample preparation to ensure complete reduction. Load a greater amount of protein to.....

• • How to Create a Protein Interaction Network if Blueberry Is Not Available in the STRING Database

  The STRING database is a widely used resource for protein-protein interaction (PPI) information, but it only covers a subset of known biological species. If the target species, such as blueberry, is not represented in STRING, the following strategies can be employed to construct a protein interaction network: 1. Prediction Based on Homology If closely related species have documented protein-protein interactions in STRING or other databases, these data can be utilized to infer potential interactions in......

• • How to Detect Target Proteins by Western Blot

  Western blotting quantitatively analyzes the expression levels of target proteins by separating protein samples through SDS-PAGE, transferring them to membranes, and subsequently binding and detecting with specific antibodies. The general procedure includes sample preparation, protein quantification, electrophoretic separation, membrane transfer, antibody binding, and detection. Detailed steps and important considerations are as follows: 1. Sample Preparation Collect cell or tissue samples and lyse them....

• • What is Chemical Proteomics

  Chemical proteomics is an interdisciplinary research area that integrates chemical biology with proteomic approaches to identify and investigate interactions between small molecules and proteins. These interactions are essential to numerous biological processes, particularly in the context of drug discovery and development. Main Objectives 1. Protein Identification and Quantification Proteins are identified and quantified using chemical strategies to elucidate their expression patterns and regulatory ......

• • What Key Points Should Be Noted in Western Blot (WB) Experiments

  In Western blot (WB) experiments, it is essential to carefully address several critical operational factors to ensure experimental accuracy and reproducibility. The main considerations are outlined as follows: Sample Preparation 1. Optimal Sample Amount An excessive or insufficient amount of sample can compromise the accuracy of experimental outcomes. 2. Application of Protease Inhibitors Protease inhibitors should be incorporated during sample preparation to prevent protein degradation. 3. Extraction......

• • Why Use Immunoprecipitation to Detect the Target Protein

  Western Blot (WB) and Immunoprecipitation (IP) are two distinct techniques, each characterized by specific applications and advantages. While both rely on antibodies, they serve different purposes and provide distinct types of information. Western Blot (WB) 1. Main Purpose To identify the presence of a specific protein within a sample and to estimate its relative abundance. 2. Information Provided Protein molecular weight, expression level, and potential post-translational modifications.......

• • What Are the Common Types of Data in Mass Spectrometry

  Mass spectrometry (MS) is an analytical technique that provides compositional information about a sample by measuring the mass-to-charge ratio (m/z) and the relative abundance of ions. Commonly encountered types of mass spectrometry data include: 1. Mass Spectrum A mass spectrum is a graphical representation in which the mass-to-charge ratio (m/z) of ions is plotted along the x-axis, while their relative abundance is plotted along the y-axis. This spectrum visually represents the distribution of ions ......

• • Why Is SDS-PAGE Not a Straight Line? Protein Precipitation with Ethanol and Redissolution with Ammonium Bicarbonate

  SDS-PAGE is a widely used technique for separating proteins according to their molecular weight. If the resulting bands do not form a straight line, several factors might be responsible: 1. Overloading or Underloading of Sample Excessive sample loading can lead to smeared or overlapping bands, while insufficient loading may result in faint or smeared bands. It is recommended to optimize the sample loading amount to an appropriate level. 2. Improper Sample Pretreatment Prior to SDS-PAGE, samples typically...

• • Can Protein Samples in Gels Be Stored Long-Term, Shipped Safely, and Used for Mass Spectrometry

  Protein samples embedded in gels can be stored long-term under appropriate conditions; however, the following considerations are important: 1. Storage Conditions Gel slices should be stored at 4°C or lower to minimize protein degradation. They should be tightly sealed with laboratory-grade plastic film, and moistened paper towels should be placed around the gel to prevent drying. Additionally, soaking the gel slices in a buffer containing 50% glycerol may further help retain moisture and prevent desiccation