416 Part III: Muscle Foods
protein spots may be used to distinguish closely
related species.
From early on, proteomic methods have been rec-
ognized as a potential method of fish species identi-
fication. During the 1960s one-dimensional electro-
phoretic techniques were developed to identify the
raw flesh of various species (Cowie 1968, Mackie
1969, Tsuyuki et al. 1966); this was soon followed
by methods to identify species in processed or
cooked products (Mackie 1972, Mackie and Taylor
1972). These early efforts were reviewed in 1980
(Hume and Mackie 1980, Mackie 1980).
More recently, 2DE-based methods have been
developed to distinguish various closely related
species, such as the gadoids or several flat fishes
(Piñeiro et al. 1999, Piñeiro et al. 1998, Piñeiro et al.
2001). Piñeiro et al. have found that Cape hake
(Merluccius capensis)and European hake (Merluc-
cius merluccius)can be distinguished on 2D gels
from other closely related species by the presence of
a particular protein spot that they identified, using
nanoelectrospray ionization mass spectrometry, as
nucleoside diphosphate kinase (Piñeiro et al. 2001).
Lopez et al., studying three species of European
mussels, Mytilus edulis, Mytilus galloprovincialis
and Mytilus trossulus, found that M. trossuluscould
be distinguished from the other two species on foot
extract 2D gels by a difference in a tropomyosin
spot. They found the difference to be due to a single
T to D amino acid substitution (Lopez et al. 2002).
Recently, Martinez and Jakobsen Friis went further
and attempted to identify not only the species pres-
ent, but also their relative ratios in mixtures of sever-
al fish species and muscle types (Martinez and
Jakobsen Friis 2004). They concluded that such a
strategy would become viable once a suitable num-
ber of markers have been identified, although detec-
tion of species present in very different ratios is
problematic.
IDENTIFICATION AND
CHARACTERIZATION OF
ALLERGENS
Food safety is a matter of increasing concern to food
producers and should be included in any considera-
tion of product quality. Among issues within this
field that are of particular concern to the seafood
producer is that of allergenic potential. Allergic re-
actions to seafood affect a significant part of the
population: about 0.5% of young adults are allergic
to shrimp (Woods et al. 2002). Seafood allergies are
caused by an immunoglobulin E–mediated response
to particular proteins, including structural proteins
such as tropomyosin (Lehrer et al. 2003). Proteo-
mics provide a highly versatile toolkit to identify
and characterize allergens. As yet, these have seen
little use in the study of seafood allergies, although
an interesting and elegant approach has been report-
ed by Yu et al. (Yu et al. 2003a) at National Taiwan
University. These authors, studying the cause of
shrimp allergy in humans, performed 2DE on crude
protein extracts from the tiger prawn, Penaeus mon-
odon, blotted the 2D gel onto a polyvinyl difluoride
(PVDF) membrane, and probed the membranes with
serum from confirmed shrimp allergic patients. The
allergens were then identified by MALDI-TOF mass
spectrometry of tryptic digests. The allergen was
identified as a protein with close similarity to argi-
nine kinase. The identity was further corroborated
by cloning and sequencing the relevant cDNA. A
final proof was obtained by purifying the protein,
demonstrating that it had arginine kinase activity
and reacted to serum IgE from shrimp allergic
patients and, furthermore, induced skin reactions in
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