Protein Analysis FAQ

• • Difference Between Sequencing and Mass Spectrometry: Is Mass Spec More Limited

  Sequencing and mass spectrometry are distinct biomolecular analysis techniques, each with unique applications and advantages. Sequencing Primarily used for analyzing nucleic acid sequences (DNA or RNA), sequencing provides detailed genomic or transcriptomic information. It enables the identification of complete gene sequences, mutations, and gene expression differences, making it essential in genomics, genetics, and molecular biology research. Mass Spectrometry Mainly used to analyze proteins, metabolites..

• • What Are the Advantages of Gas Chromatography-Mass Spectrometry

  Gas Chromatography-Mass Spectrometry (GC-MS) combines the strengths of both gas chromatography (GC) and mass spectrometry (MS), offering several key advantages: 1. Strong Separation Capability GC effectively separates complex mixtures into individual compounds. By adjusting column properties and operating conditions, GC-MS achieves high-precision separation, enhancing analytical accuracy and reliability. 2. High Sensitivity MS provides ultra-high sensitivity, detecting trace-level compounds. In GC-MS, GC...

• • How Is Ubiquitinated Protein Generally Detected

  Ubiquitination is a post-translational modification process in which ubiquitin is attached to proteins, regulating their degradation, signaling, and other functions. Common methods for detecting ubiquitinated proteins include: 1. Western Blot (WB) Protein extraction is performed on cell or tissue samples, followed by SDS-PAGE separation. The separated proteins are transferred to a membrane and incubated with an anti-ubiquitin antibody. If ubiquitination occurs, higher molecular weight ubiquitinated ........

• • How to Predict Its Glycosylation Sites If Only an Amino Acid Sequence Is Known

  Protein site modifications can indeed cause molecular weight changes, especially upward shifts. Predicting glycosylation sites in a protein typically involves identifying which amino acid residues might be modified by carbohydrate molecules. The most common types of glycosylation are N-linked glycosylation and O-linked glycosylation. Given a known partial amino acid sequence, glycosylation sites can be predicted using the following methods: Sequence-Based Rules 1. For N-linked glycosylation, a typical......

• • Does Modification of Protein Sites Cause an Increase in Molecular Weight

  Protein site modifications can indeed alter molecular weight, often leading to an upward shift (increase). Protein modifications are common biological phenomena that regulate various functions, including protein activity, stability, and localization. These modifications typically involve the addition of small molecular groups, such as phosphorylation, ubiquitination, glycosylation, and lipid anchoring. Since these added molecules have their own molecular weight, modified proteins usually exhibit an ........

• • When Performing a WB Experiment, Why Is It Necessary to Add Proteases and Inhibitors When Extracting Proteins

  During a Western Blot (WB) experiment, the addition of proteases and inhibitors during the protein extraction process is necessary for the following reasons: Addition of Proteases 1. The presence of proteases may degrade the target proteins; therefore, adding proteases during protein extraction can prevent protein degradation. 2. Proteases can degrade intracellular proteins, including receptor proteins on the cell membrane, cytoskeletal proteins, etc. By adding proteases, these intracellular proteins can...

• • How to Remove the Solvent Peak in Gas Chromatography So That It Does Not Display in the Chromatogram

  In gas chromatography (GC) analysis, the presence of a solvent peak may interfere with the detection of target compounds. Several strategies can be employed to mitigate or eliminate solvent peak interference: 1. Extending Retention Time Modifying chromatographic conditions, such as reducing the column temperature or utilizing a longer chromatographic column, can increase the retention time of target compounds. This facilitates better separation between target compound peaks and the solvent peak ......

• • Will the Protein Sample Degrade if Left at Room Temperature Overnight

  Protein stability is primarily influenced by the specific protein, sample composition, and storage conditions. However, for most proteins, storage at room temperature overnight can result in degradation. For critical protein samples, freezing immediately following purification and storing at -80°C is advisable. Alternatively, adding suitable stabilizers or protease inhibitors and storing at 4°C for short-term preservation can help maintain stability. For long-term storage, samples should be kept at -20°C...

• • What Software Is Used for Protein Mass Spectrometry? Why Is Protein Identification Yield Low

  In protein mass spectrometry analysis, a variety of software tools are available for qualitative protein identification. Below are several commonly used platforms: 1. Mascot A widely adopted commercial tool developed by Matrix Science. It supports data interpretation from various mass spectrometry techniques by performing database searches across multiple protein repositories. 2. SEQUEST Originally developed by John Yates III and currently distributed by Thermo Fisher Scientific, SEQUEST is typically ......

• • Why Does Loading Too Large a Sample Volume in Gel Filtration Chromatography Lead to Peak Overlapping

  Gel filtration chromatography (also referred to as gel permeation chromatography or size-exclusion chromatography) is a widely used biochemical technique for separating molecules based on size. When an excessively large sample volume is loaded, peak overlapping may occur. This phenomenon can be attributed to the following factors: 1. Sample Distribution In gel filtration chromatography, molecules are separated according to their hydrodynamic size as they pass through the gel matrix. Larger molecules are....