Chapter 9
Human Blood Plasma Investigation Employing 2D
UPLC-UDMS
E Data-Independent Acquisition Proteomics
Licia C. Silva-Costa, Bradley J. Smith, Pamela T. Carlson,
Gustavo H. M. F. Souza, and Daniel Martins-de-Souza
Abstract
Proteomic tools are especially useful when it comes to investigating complex samples such as human blood
plasma, in which protein quantities can span across up to ten orders of magnitude. Ultra definition mass
spectrometry, in combination with two-dimensional liquid chromatography, provides better coverage of
complex proteomes and allows for better control of collision energy, keeping the fragmentation benefits of
high collision energy associated with drift time measurements from ion mobility separation. Here, we
present a protocol to assist in the identification of proteins in human blood plasma and other similar samples
with a large dynamic range.
Key words Multifactorial disorders, Immunoaffinity depletion, Tryptic digestion, Serum, Depletion,
Mass spectrometry
1 Introduction
Several human diseases are considered “multifactorial,” given the
complex interaction of genes, their direct (RNA) and indirect products (proteins, including their modifications and metabolites),
and the environment. Such complexity makes multifactorial diseases challenging for researchers and physicians. Understanding
how biological mechanisms and changes in gene expression are
implicated in these diseases is a long journey since it is imperative
to bring together data and analyses from multiple areas and fields of
study such as genetics, epigenetics, analytical chemistry, statistics,
and mathematical models, among many others. Due to the complex nature of multifactorial diseases, the use of personalized medicine is becoming more important in the research and medical fields.
Overall, goals of personalized medicine include the development of
reliable prediction models of individual risk as well as knowledge
about therapeutic outcomes [1, 2]. Other goals include
Mo ´ nica Carrera and Jesu ´ s Mateos (eds.), Shotgun Proteomics: Methods and Protocols, Methods in Molecular Biology, vol. 2259,
https://doi.org/10.1007/978-1-0716-1178-4_9, © Springer Science+Business Media, LLC, part of Springer Nature 2021
153
Human Blood Plasma Investigation Employing 2D
UPLC-UDMS
E Data-Independent Acquisition Proteomics
Licia C. Silva-Costa, Bradley J. Smith, Pamela T. Carlson,
Gustavo H. M. F. Souza, and Daniel Martins-de-Souza
Abstract
Proteomic tools are especially useful when it comes to investigating complex samples such as human blood
plasma, in which protein quantities can span across up to ten orders of magnitude. Ultra definition mass
spectrometry, in combination with two-dimensional liquid chromatography, provides better coverage of
complex proteomes and allows for better control of collision energy, keeping the fragmentation benefits of
high collision energy associated with drift time measurements from ion mobility separation. Here, we
present a protocol to assist in the identification of proteins in human blood plasma and other similar samples
with a large dynamic range.
Key words Multifactorial disorders, Immunoaffinity depletion, Tryptic digestion, Serum, Depletion,
Mass spectrometry
1 Introduction
Several human diseases are considered “multifactorial,” given the
complex interaction of genes, their direct (RNA) and indirect products (proteins, including their modifications and metabolites),
and the environment. Such complexity makes multifactorial diseases challenging for researchers and physicians. Understanding
how biological mechanisms and changes in gene expression are
implicated in these diseases is a long journey since it is imperative
to bring together data and analyses from multiple areas and fields of
study such as genetics, epigenetics, analytical chemistry, statistics,
and mathematical models, among many others. Due to the complex nature of multifactorial diseases, the use of personalized medicine is becoming more important in the research and medical fields.
Overall, goals of personalized medicine include the development of
reliable prediction models of individual risk as well as knowledge
about therapeutic outcomes [1, 2]. Other goals include
Mo ´ nica Carrera and Jesu ´ s Mateos (eds.), Shotgun Proteomics: Methods and Protocols, Methods in Molecular Biology, vol. 2259,
https://doi.org/10.1007/978-1-0716-1178-4_9, © Springer Science+Business Media, LLC, part of Springer Nature 2021
153