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Background:
Anisakis simplex is
a nematode which can
parasitize humans,
producing
anisakiasis and can
induce
immunoglobulin-(Ig)-
Emediated allergic
symptoms. Parasite
recombinant proteins,
such as the major
allergen Ani s 1,
may be useful tools
to avoid
misdiagnosis of A
simplex allergy due
to cross-reactivity
when whole parasite
extracts are used.
Objective: To
obtain Ani s 1
allergen as a
recombinant protein
with IgE-binding
properties similar
to its natural
counterpart.
Methods: Ani
s 1-encoding cDNA
was amplifi ed by
polymerase chain
reaction and cloned.
The allergen was
expressed in
Escherichia coli as
a nonfusion protein.
Natural and
recombinant Ani s 1
were investigated by
means of Western
blotting, enzyme
allergosorbent test,
enzyme-linked
immunosorbent assay
(ELISA), and ELISA
inhibition using
sera from 53
patients with A
simplex allergy.
Results:
Residues of the
amino acid sequence
of the encoded
protein were 99.4%
identical to the
reported one. Purifi
ed rAni s 1 was
obtained with a
yield of 2 mg/L of
culture while the
yield of the natural
counterpart was only
50 μ/g of larvae.
rAni s 1 reactivity
was not
significantly
different from that
of the natural
allergen; the
correlation was
excellent (ρ = 0.92,
P < .001). ELISA-inhibition
experiments showed
that the dose-response
inhibition curve
obtained with rAni s
1 overlapped with
that of nAni s 1. In
an enzyme
allergosorbent
analysis, 86.8% of
the A simplex-allergic
patient sera reacted
to rAni s 1.
Conclusion:
Recombinant Ani s 1
is immunochemically
equivalent to its
natural counterpart
and therefore might
be useful for the in
vitro diagnosis of
anisakiasis and A
simplex-mediated
allergy.
Key words:
Nematode.
Recombinant allergen.
Biologically active
allergen Diagnostic
tool. Major allergen. |
