References/Publications for Mass spectrometry (MS) of Fra e 2

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19886925 Allergy. 2010 May;65(5):571-80. doi: 10.1111/j.1398-9995.2009.02231.x. Epub 2009
Nov 3.

Evaluation of ash pollen sensitization pattern using proteomic approach with
individual sera from allergic patients.

Poncet P(1), Senechal H, Clement G, Purohit A, Sutra JP, Desvaux FX, Wal JM,
Pauli G, Peltre G, Gougeon ML.


25219108 J Investig Allergol Clin Immunol. 2014;24(4):257-66.

Characterization of profilin and polcalcin panallergens from ash pollen.

Mas S, Garrido-Arandia M, Batanero E, Purohit A, Pauli G, Rodríguez R, Barderas
R, Villalba M.

BACKGROUND: Ash (Fraxinus excelsior) is an important source of allergenic pollen
in temperate areas of Europe. Profilin and polcalcin are 2 important panallergens
involved in cross-reactivity between different sources.
OBJECTIVE: To clone and produce Fra e 2 (profilin) and Fra e 3 (polcalcin) as
recombinant proteins and evaluate their immunological properties using the
natural forms obtained from ash pollen.
METHODS: Total RNA from ash pollen was used as a template to obtain the specific
complementary DNA (cDNA) sequences of the 2 panallergens. The cDNA-encoding
sequences were cloned into the pET11b expression vector and used to transform
BL21 (DE3) Escherichia coli cells. Proteins were expressed, purified by
chromatography, and characterized structurally by circular dichroism, mass
spectrometry, and immunologically by western blot and ELISA using profilin and
polcalcin polyclonal antibodies and human sera from ash pollen-sensitized
patients.
RESULTS: Profilin and polcalcin amino acid sequences from ash pollen showed a
high degree of identity with homologous allergens from different sources. The
cDNA-encoding allergen sequences were expressed as nonfusion recombinant proteins
and purified to homogeneity. Secondary structure values were similar to those
obtained from other members of these families. Allergenic properties of the
recombinant allergens were observed to be equivalent to those of the natural
counterparts of F excelsior pollen.
CONCLUSIONS: Fra e 2 and Fra e 3 recombinant allergens might be used in clinical
diagnosis to determine profilin- and polcalcin-specific IgE levels present in the
sera of ash pollen-sensitized patients, thus facilitating the finding of the
sensitizing source in areas with complex sensitization profiles.


PMID: 25219108 [Indexed for MEDLINE]