fractionation alters the relative abundance of proteins by a factor of 10 to 100...”^7
At the other hand, other approaches show that proper sample preparation is the first,
key step for quantitative detection of some proteins, e.g., high glycosylated ones
and proteins that contain hydrophobic transmembrane domains. Additionally,
proteolytic digestion as the second, not less important step in sample preparation
is frequently neglected, as well as the simple fact that a peptide that is not generated
by proteolysis consequently cannot be detected by any of high-performance, up-to-
date mass spectrometer.^8 As already pointed above, each step in omic investi-
gations (and in proteomic ones as a part of this complex) has to be clearly defined
in translational communication between the clinician and basic researcher.
The recent progress report of the Human Proteome Project (HPP) for the years
2013–2014 that summarizes the work of about 50 teams worldwide shows that there
are still 3844 “missing” proteins to be found out of 19,490 proteins coded by
corresponding genes. Currently, these proteins have no or only inadequate docu-
mentation and are still waiting for their validation.^9 Strategies for searching for
missing proteins and their tissue distribution, appearance in early stage of life, their
change under special stress conditions, and other perturbations were also discussed,
as well as optimization and development of new analytical methods for the detec-
tion of low abundance proteins, proteins with one or more transmembrane helical
structures that contain sequences that cannot be cleaved by trypsin and proteins
with posttranslational modifications (PTMs). All these proteins are still difficult to
detect even if high-sensitivity mass spectrometers are used for analysis. Farrah
et al.^10 also point out that major data sets for liver, muscle, and kidney are not
present in HPP Atlas. It is well known that even these tissues contain very hydro-
phobic and very basic proteins, as well as membrane proteins with complex PTMs.
These organs are also frequently hit by diseases or damage, especially in immune
deficient and aged individuals.
Proteins in body fluids and of both cell surface and intracellular matrix are the
first ones that show detectable changes during early development of pathologic
processes in the organism and during a patient’s recovery. It is the reason that these
proteins are main targets for biomarker discovery for early disease detection, as
well as for control of healing process.^11 According to the National Cancer Institute
(NCI), a biomarker is defined as “a biological molecule found in blood, other body
fluids or tissues that is a sign of normal or abnormal processes or of a condition of
disease.”^12 A biomarker is a molecule that is present in both normal and patho-
logical populations, but with strongly altered levels and molecular forms in affected
patients. This molecule can be also newly expressed or altered in tissues and body
(^7) Mann et al. ( 2013 ), pp. 582–590.
(^8) Cao et al. ( 2013 ), pp. 8112–8120.
(^9) Lane et al. ( 2014 ), pp. 15–20.
(^10) Farrah et al. ( 2013 ), pp. 162–171.
(^11) Goldknopf ( 2008 ), pp. 1–8.
(^12) Boschetti et al. ( 2012 ), pp. 22–41.
182 D. Josic ́and U. Andjelkovic ́