Protein Analysis FAQ

• • What Concentration is Corresponding When S/N Ratio is 3 in Chromatographic System Suitability Testing by HPLC

  In high-performance liquid chromatography (HPLC) system suitability testing, when the signal-to-noise ratio (S/N) is 3, the corresponding concentration is typically regarded as the method's limit of detection (LOD). The signal-to-noise ratio (S/N) serves as an indicator of the analytical method's sensitivity, assessing the relationship between the signal of the target component and the background noise. A S/N ratio of 3 suggests that the signal from the target component is sufficiently strong to be ........

• • How to Estimate the Protein Concentration in a Band Based on Protein Markers

  Estimating protein concentration in SDS-PAGE bands using protein markers (also known as protein molecular weight standards or ladders) is commonly performed by comparing the intensity of staining in sample bands to that of marker bands with known protein concentrations. However, this approach provides only an approximate estimation. The general procedure is as follows: 1. Running SDS-PAGE Load the protein marker and your sample into separate lanes of an SDS-PAGE gel and run electrophoresis. Ensure that.....

• • Native Polyacrylamide Gel Electrophoresis for Proteins

  Native polyacrylamide gel electrophoresis (Native PAGE) is an electrophoretic technique for separating proteins under non-denaturing conditions. Unlike SDS-PAGE, Native PAGE preserves the native three-dimensional structure and biological function of proteins. Principle Native PAGE operates by applying an electric field to induce the migration of proteins through a polyacrylamide gel. The mobility of proteins in the gel is determined by their net charge, molecular size, and conformation.

• • How Electrophoresis Separates and Purifies Proteins Based on Their Physicochemical Properties

  Electrophoresis is a technique that separates and purifies proteins and other biomolecules by exploiting their migration in an electric field. The principle behind this technique is based on several key physicochemical properties of proteins: 1. Protein Charge Characteristics Proteins are composed of amino acid residues, each of which typically carries a charge. Some amino acids have a positive charge, while others possess a negative charge. At a specific pH, a protein will acquire a net overall charge.....

• • Common Types of Proteins Used in WB Experimental Protein Extraction

  When performing protein extraction for WB experiments, the commonly extracted protein fractions include the following: Total Cellular Proteins This is the most commonly extracted protein fraction, obtained by lysing both the plasma and nuclear membranes to release all intracellular proteins. This extraction approach is widely used to analyze overall protein expression in cells. Nuclear Proteins Nuclear proteins are isolated by selectively lysing the cytoplasmic membrane while preserving nuclear integrity...

• • What Are the Methods for Determining the Molecular Weight Distribution of Polymers

  Molecular weight distribution (MWD) of polymers describes the relative proportions of polymer molecules with different molecular weights within a given sample. Determining MWD is crucial for understanding the structure and properties of polymeric materials. The following are some commonly used methods for analyzing the molecular weight distribution of polymers: 1. Gel Permeation Chromatography (GPC/SEC) Gel permeation chromatography (also known as size-exclusion chromatography, SEC) is the most widely......

• • What Should Be Done if the Flow Rate Is Too Slow During Protein Purification Through a Nickel Column

  Nickel columns (Ni-NTA) are a widely used method for protein purification, utilizing the binding of His-tagged proteins to six consecutive histidine residues (His-tag). If you experience a slow flow rate while purifying proteins using a nickel column, you can try the following approaches: Check the Piston and Connections Ensure that the piston and connections are correctly installed and undamaged. If any issues are found, replace the piston or connections as needed. Reduce the Sample Concentration .........

• • What Determines the Migration Direction of Proteins in an Electric Field

  In different types of electrophoresis, the migration direction of proteins is determined by their charge state as well as the electrophoretic conditions and medium used. Below are several common types of electrophoresis and the factors determining protein migration direction: 1. Native PAGE (Non-denaturing Electrophoresis) In native PAGE, proteins maintain their natural state and are not denatured. The migration direction of proteins depends on their charge state at the pH of the electrophoresis buffer.....

• • Is the Sample Preparation for Protein Detection Using MALDI-TOF and QE Mass Spectrometry the Same

  MALDI-TOF (Matrix-Assisted Laser Desorption/Ionization Time-of-Flight) mass spectrometry and QE (Quadrupole-Orbitrap) mass spectrometry are two different mass spectrometry techniques, and their pretreatment steps in protein detection have some differences. The pretreatment for MALDI-TOF tends to involve directly mixing the sample with a matrix to form crystals, whereas the QE mass spectrometry pretreatment focuses more on liquid chromatography separation and ionization of proteins or peptides.

• • Why Are There So Many Bubbles on the Edge of the Western Blot Gel

  The formation of bubbles along the edges of Western blot gels can result from several factors, including the following: 1. Bubbles Introduced During Gel Preparation During the preparation of polyacrylamide gels, inadequate mixing or uneven stirring of the gel solution can lead to bubble formation. To minimize this, the gel solution should be thoroughly mixed and then allowed to stand for a period before use, facilitating the escape of trapped air. 2. Bubbles Formed During Gel Pouring If the gel solution....