Production and evaluation of the recombinant antigen TES-30 of Toxocara canis for the immunodiagnosis of toxocariasis.
BACKGROUND
Toxocara canis is a pathogenic nematode of canines which can be accidentally transmitted to humans. Although serology is the most important diagnostic tool for this zoonosis, diagnostic kits use crude excretion/secretion antigens, most of them being glycoproteins which are not species-specific and may cross-react with antibodies generated against other parasites.


OBJECTIVE
To produce the rTES-30 recombinant antigen of Toxocara canis and evaluate it in the immunodiagnosis of toxocariasis.


METHODS
The gene that codes for TES-30 was cloned in the expression vector pET28a (+) using single-stranded oligonucleotides united by PCR. The protein rTES-30 was purified by Ni2+ affinity chromotography. Seroreactivity of rTES-30 was evaluated by immunoblot. Given that there is no gold standard test, the behaviour of the antigen was compared with the method that is routinely used to immunodiagnose toxocariasis, i.e., the conventional ELISA technique using excretion/secretion antigens.


RESULTS
The rTES-30 was produced from an Escherichia coli LB culture which yielded 2.25 mg/L of the antigen with a purity of 95%. The results obtained showed 73% (46/63) concordance of reactivity between the rTES-30 immunoblot and the conventional ELISA, and 100% concordance with the nonreactive sera (21). Nineteen of the 21 sera positive for other parasitoses reacted with ELISA, while only seven of these were positive with the rTES-30 immunoblot. Concordance between the ELISA and the immunoblot was moderate (kappa coefficient: 0.575; 95% CI: 0.41- 0.74).


CONCLUSIONS
The data presented show the potential of the rTES-30 inmunoblot for confirmation of possible ELISA positives, not only in epidemiological studies, but also as a candidate for the development of diagnostic tests for ocular toxocariasis in Colombia.
Grupo de Parasitología, Facultad de Medicina, Universidad de Antioquia, Medellín, Colombia. biomedica@ins.gov.co.
Ana M
AM
Olave
Jairo A
JA
Mesa
Jorge H
JH
Botero
Edwin B
EB
Patiño
Gisela M
GM
García
Juan F
JF
Alzate
D000818:Animals; D000947:Antigens, Helminth / Q000097:blood* / Q000235:genetics / Q000302:isolation & purification; D001483:Base Sequence; D002846:Chromatography, Affinity; D004797:Enzyme-Linked Immunosorbent Assay; D004926:Escherichia coli; D015822:Eye Infections, Parasitic / Q000175:diagnosis; D005813:Genes, Synthetic; D006801:Humans; D015151:Immunoblotting; D010446:Peptide Fragments / Q000032:analysis / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification; D011993:Recombinant Fusion Proteins / Q000032:analysis / Q000302:isolation & purification; D012995:Solubility; D017166:Toxocara canis / Q000276:immunology*; D014120:Toxocariasis / Q000097:blood / Q000175:diagnosis*
D000947:Antigens, Helminth; D010446:Peptide Fragments; D011993:Recombinant Fusion Proteins
Toxocara canis; antigen; immunologic tests.; toxocariasis/diagnosis