Protein Analysis FAQ

• • What Are the Causes of Negative Integrated Peak Area in High Performance Liquid Chromatography

  In High Performance Liquid Chromatography (HPLC), the integrated peak area should always be positive, as it reflects the quantity of a compound represented by the area under the chromatographic peak. A negative integrated peak area may arise due to the following reasons: 1. Excessively Broad Peak If a chromatographic peak is too broad, it may result in a negative integrated peak area. This can occur when the peak extends significantly beyond the baseline estimation region, causing the integration ..........

• • How Is a Polypeptide Defined? What Amino Acid Length Is Considered a Polypeptide

  Polypeptides are biological macromolecules composed of several to hundreds of amino acids linked by peptide bonds. They serve as the fundamental building blocks of proteins. While there is no universally accepted definition based strictly on amino acid length, polypeptides and proteins are generally distinguished by the number of amino acid residues they contain. Typically, macromolecules consisting of 50 or fewer amino acids are referred to as polypeptides. This distinction is primarily based on two main..

• • What Is the Maximum Temperature That Can Be Set for the Injection Port in Gas Chromatography-Tandem Mass Spectrometry

  The injection port temperature in gas chromatography-tandem mass spectrometry (GC-MS/MS) is primarily determined by the instrument's design and the specifications provided by the manufacturer. The allowable temperature range of the injection port may vary depending on the specific model of the gas chromatographic system. In general, the injection port temperature is adjustable within a defined range, with most commercial systems permitting settings between 40 °C and 350 °C.

• • What Are the Applications of Proteomics in the Field of Biology

  Proteomics, the large-scale study of all proteins within a biological system, represents a critical branch of biology, medicine, drug development, and other related disciplines. In biology, proteomics has been widely applied in the following areas: 1. Protein Identification Proteomics enables the identification of protein abundance, types, and post-translational modifications. Using mass spectrometry, researchers can analyze complex protein mixtures to determine amino acid sequences, structural ........

• • How to Address Internal Standard Detection Failure in Positive Ion Mode During LC-MS Quantification

  In liquid chromatography-mass spectrometry (LC-MS) analysis, if an internal standard is detectable in negative ion mode but fails to appear in positive ion mode, the following factors may be responsible: 1. Chemical Properties of the Internal Standard The internal standard may not undergo efficient ionization under positive ion mode conditions. Consider selecting an alternative internal standard that is more amenable to ionization in positive ion mode. 2. Ion Source Conditions Adjust the ion source ........

• • Is Protein Expression in Phosphoproteomics Related to Phosphorylation Level or Protein Activity

  Phosphoproteomics is a technique for investigating protein phosphorylation modifications, providing insights into cellular signaling, regulatory mechanisms, and protein function. In phosphoproteomics, protein expression typically refers to the total abundance of a given protein, whereas phosphorylation level denotes the relative occupancy of phosphorylation sites within the protein. Protein activity, defined as the functional capacity of the protein in vivo, may be modulated by phosphorylation events.

• • How to Analyze Proteomic Mass Spectrometry with MaxQuant

  MaxQuant is a widely used software platform for the analysis of proteomic mass spectrometry data. It is designed to process bottom-up proteomic data, enabling both protein identification and quantification. The following outlines the essential steps for analyzing proteomic mass spectrometry data using MaxQuant: 1. Prepare Data and Software Ensure that the latest version of MaxQuant is installed (available at: https://www.maxquant.org/). Prepare the raw mass spectrometry data files (typically in .raw format)

• • Statistical Analysis of Differentially Expressed Proteins: How to Know Which One is the Most Significant

  In the statistical analysis of differentially expressed proteins, various statistical methods are employed to compare protein expression levels across different samples and to identify proteins exhibiting significant differences. Commonly used approaches include the t-test, analysis of variance (ANOVA), and the Wilcoxon rank-sum test. These methods yield a significance level (P-value) for each protein, with smaller P-values indicating a higher degree of statistical significance. Typically, proteins with....

• • What Are the Applications of Hydrophilic Interaction Liquid Chromatography in Proteomics

  Hydrophilic Interaction Liquid Chromatography (HILIC), a variant of High-Performance Liquid Chromatography (HPLC), is primarily employed for the separation of polar compounds. Owing to its unique separation mechanism, HILIC has emerged as a powerful tool in proteomic analysis and the investigation of complex biological samples. In the context of proteomics, HILIC finds application in the following key areas: 1. Peptide Enrichment HILIC enables efficient enrichment and separation of polar peptides within....

• • Can Unpurified Samples Be Used in Protein CD Analysis? What Are the Protein Concentration Requirements

  Can Unpurified Samples Be Used in Protein Circular Dichroism (CD) Analysis? In protein CD analysis, the use of unpurified samples is generally not recommended. Such samples may contain various contaminants or non-target components that can interfere with the CD spectra, resulting in ambiguous or difficult-to-interpret signals. To obtain accurate and reliable data, it is advisable to purify the protein beforehand to ensure that the sample primarily consists of the target protein. Are There Specific .........