Biopharmaceuticals Biotechnology and Protein Analysis

Determination of Very Low Abundance Diagnostic Proteins in Serum Using Immunocapture LC–MS/MS

July 01, 2017

ByTrine Grønhaug Halvorsen|Léon Reubsaet

Special Issues

There is growing interest in the determination of endogenous proteins in biological samples for diagnostic purposes, because a concentration increase or decrease of such proteins can allows us to monitor the state of a pathological condition such as cancer. Immunocapture LC–MS/MS analysis combines the workflow of conventional immunological assays with LC–MS analysis. This article describes typical challenges, such as cross reactivity and the mass spectrometer’s dynamic range, as well as the advantages of isoform differentiation and multiplexing.

A New Mass Spectrometry Method for Protein Analysis

March 22, 2017

Proteomics and structural biology require specialized mass spectrometry methods for characterizing protein structures and conformations. Jennifer S. Brodbelt, a professor of chemistry at the University of Texas at Austin, focuses on the development and application of photodissociation mass spectrometry for studying biological molecules such as peptides, proteins, nucleic acids, oligosaccharides, and lipids. She recently spoke with Spectroscopy about her work with this technique. She is the winner of the 2017 ANACHEM Award, which will be presented at the SciX meeting in October 2017. The award is presented annually to an outstanding analytical chemist based on activities in teaching, research, administration, or other activities that have advanced the art and science of the field.

Absolute Quantification of Proteins and Peptides by ICP-MS

November 01, 2016

ByArnaud Delobel|Géry Van Vyncht|Philippe De Raeve|Juliusz Bianga

Special Issues

Despite all of the recent advances in analytical technologies dedicated to biotherapeutics, accurate protein quantification remains a challenge for the biopharmaceutical industry. UV spectrophotometry is commonly used for batch testing, but it requires the knowledge of the extinction coefficient of the protein, whose experimental determination requires the accurate concentration of a reference standard obtained by an absolute quantification method. To address the need for a fast analytical method capable of accurately quantifying a protein without any specific reference substance, an isotope dilution ICP-MS method was developed and validated, based on sulfur determination, allowing very accurate determination of a single protein in solution after microwave digestion.

Protein Secondary Structure Determination Using Drop Coat Deposition Confocal Raman Spectroscopy

October 01, 2016

ByJeremy Peters|Eugene Park|Ravi Kalyanaraman|Anna Luczak|Varsha Ganesh

Spectroscopy

The accurate determination of protein structure is integral to the medical and pharmaceutical communities’ ability to understand disease, and develop drugs. Current techniques (CD, IR, Raman) for protein structure prediction provide results that can be poorly resolved, while high resolution techniques (NMR, X-ray crystallography) can be both costly and time-consuming. This work proposes the use of drop coat deposition confocal Raman spectroscopy (DCDCR), coupled with peak fitting of the Amide I spectral region (1620–1720 cm-1) for the accurate determination of protein secondary structure. Studies conducted on BSA and ovalbumin show that the predictions of secondary structure content within 1% of representative crystal structure data is possible for model proteins. The results clearly demonstrate that DCDCR has the potential to be effectively used to obtain accurate secondary structure distributions for proteins.

Advances in ICP-MS Detection of Selenium in Proteins

February 01, 2016

Spectroscopy

Joanna Szpunar of the National Research Council of France discusses her use of ICP-MS methods to detect trace levels of human selenoproteins in cell extracts and for the identification of selenium containing proteins in rice.

Advances in ICP-MS Detection of Selenium in Proteins

March 20, 2015

Selenoproteins play an important role in human physiology and health, and as a result, sensitive methods are needed for their analysis. Joanna Szpunar of the National Research Council of France has been working with bioinorganic speciation analysis and hyphenated techniques for metallomics studies for some time. She recently spoke to Spectroscopy about using laser-ablation inductively coupled plasma–mass spectrometry (LA-ICP-MS) to detect trace levels of human selenoproteins in cell extracts and about her work with ICP-MS-assisted electrospray tandem MS for the identification of selenium-containing proteins in rice grown on seleniferous soils.

UV Resonance Raman Spectroscopy Analysis of Protein Structure and Folding

March 16, 2015

UV resonance Raman spectroscopy examines how UV light interacts with the electrons of samples and provides information about their molecular structure and dynamics. Sanford A. Asher, Distinguished Professor of Chemistry at the University of Pittsburgh, is using the technique to study peptide excited states and conformations and protein folding, with the ultimate goal of helping to advance research into the mechanisms of disease. He recently spoke to us about this work.

Large-Scale Targeted Protein Quantification Using Wide Selected-Ion Monitoring Data-Independent Acquisition

October 01, 2014

Special Issues

This article describes the development of a new data-independent acquisition (DIA) workflow for protein quantification that uses a mass spectrometer that combines three types of mass analyzers to achieve lower limits of detection (LOD), higher sensitivity, more accurate quantitative results, wider dynamic range, and better reproducibility than existing high-resolution accurate-mass (HRAM) tandem mass spectrometry (MS-MS) DIA workflows.