Protein Analysis FAQ

• • How to Prepare Samples, Concentration, and Buffer for Circular Dichroism Analysis of Protein Secondary Structure?

  Circular Dichroism (CD) is a commonly used spectroscopic technique for analyzing protein secondary structure. The preparation of samples and measurement conditions significantly impact the quality and accuracy of the CD spectra. When preparing samples, pay attention to the following:   Sample Preparation 1. Protein needs to be purified, avoiding impurities and other interfering substances.   2. If the protein needs to be refolded, ensure that the correctly folded protein is obtained.   3. Avoid using ......

• • What Are the Reasons for No Peaks in Liquid Chromatography Samples?

  Liquid Chromatography (LC) is a common separation and analysis method used to detect and quantify components in complex samples. If no peaks appear during the LC process, several reasons may be responsible:   Instrument Malfunction Certain parts of the chromatograph, such as the pump, detector, injector, or column, may be faulty. Check that the instrument is properly turned on, running, and that all connections in the LC system are intact.   Column Efficiency Prolonged use or improper handling of the ......

• • What Are the Methods for Protein Covalent Modifications?

  Protein covalent modifications refer to the non-translational addition of specific chemical groups to a protein’s amino acid residues via chemical or enzymatic reactions, thereby altering its structure and function. This is a crucial process in cellular regulation and signal transduction. Below are several methods for protein covalent modifications:   Phosphorylation A kinase-catalyzed process that adds a phosphate group to the serine, threonine, or tyrosine residues of proteins. This important regula......

• • How to Predict a Peptide's Response Given Its Sequence?

  To predict a peptide's response, you can follow a systematic approach that includes sequence analysis, structure prediction, function prediction, experimental validation, machine learning and data mining, and literature search. The following details each step and its specific methods:   Sequence Analysis 1. Analysis of Basic Physicochemical Properties (1) Amino Acid Composition: Analyzing the amino acid composition helps understand its hydrophilicity or hydrophobicity. For example, peptides rich in hy......

• • Why Should Proteins Be Boiled Before SDS-PAGE Electrophoresis?

  SDS-PAGE (Sodium Dodecyl Sulfate-Polyacrylamide Gel Electrophoresis) is a commonly used technique for protein separation. The reasons for boiling proteins before SDS-PAGE can be explained as follows:   1. Protein Denaturation Heat and SDS work together to disrupt the three-dimensional structure of the protein, converting it into a linear form. This eliminates interactions between proteins, ensuring their migration during electrophoresis is not affected by their original conformation.   2. Binding of S......

• • What Are the Differences Between Solid-Phase and Liquid-Phase Peptide Synthesis?

  Peptide synthesis refers to the chemical process of creating peptide chains. It is typically performed using solid-phase synthesis or liquid-phase synthesis. There are significant differences between these two methods, as outlined below:   Synthesis Strategy 1. Solid-Phase Synthesis Involves fixing the first amino acid on a solid support (usually resin) and progressively attaching other amino acids to form the peptide chain. After each addition, unreacted materials and by-products are washed away befo......

• • What Are the Specific Steps for DSS Protein Crosslinking?

  "DSS" (Disuccinimidyl Suberate) is a common chemical crosslinker primarily used to study and stabilize protein-protein interactions. DSS is activated as an NHS ester at room temperature and reacts with amino groups in proteins to form covalent bonds. The specific steps for DSS protein crosslinking may vary depending on your experimental needs and conditions. Below is a general procedure that may help:   1. Protein Preparation Prepare the sample containing the target proteins. If the proteins are intra......

• • Can Data Points Be Removed If R² of the Standard Curve Is Below 0.99?

  In scientific experiments, the quality of the standard curve is an important indicator for evaluating the accuracy and reliability of experimental results. If the R² value of a standard curve is below 0.99, it suggests that the correlation between the data and the fitted model may not be strong enough. Removing data points to improve the R² value should be approached cautiously, with the following considerations:   Impact of Removing Data Points Removing data points to improve the R² value can sometim......

• • How to Determine the Molecular Weight and Amino Acid Residues of a Protein? Is Experimental Detection Necessary?

  To determine the molecular weight and number of amino acid residues of a protein, there are several approaches and methods: querying public databases, using tools, and experimental detection.   Using Bioinformatics Tools and Databases 1. Database Search If the protein sequence is known, you can search for its molecular weight and amino acid sequence information in public protein databases, such as UniProt, NCBI's Protein Database, and PDB (Protein Data Bank). These databases typically list known prote......

• • Why Do Western Blot Bands Appear Darker at the Edges and Lighter in the Center?

  In Western Blot (WB) analysis, bands that are darker at the edges and lighter in the center are often due to sample overloading. When the amount of protein loaded exceeds the gel's capacity, proteins may not migrate evenly during electrophoresis, especially in the central region of the gel. This results in faster or further migration at the edges, making the center of the band appear lighter.   To address this issue, reduce the amount of protein loaded onto the gel, ensuring appropriate concentration ......