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VALIDATION AND ANALYSIS OF AFLATOXIN B1 BY INDIRECT COMPETITIVE ENZYME-LINKED IMMUNOSORBENT ASSAY IN URINE SAMPLES FROM CHILDREN IN LONDRINA, PARANÁ STATE, BRAZIL

Ishikawa AT;
Redondo ES;
Yamashita CRT;
Ono EYS;
Itano FH;
Souza BDO;
Ono MA;
Kawamura O;
Hirooka EY;
Itano EN

Angélica Tiemi Ishikawa

Estéfany Santos Redondo

Cassia Reika Takabayashi Yamashita

Elisabete Yurie Sataque Ono

Flávio Hiroshi Itano

Bianca Dorana de Oliveira Souza

Mario Augusto Ono

Osamu Kawamura

Elisa Yoko Hirooka

Eiko Nakagawa Itano


Keywords

Monoclonal antibodies
Fungi
Mycotoxin
Immunodiagnosis
Food security

Abstract

Aflatoxins (AFs), secondary metabolites produced by fungi of the genus Aspergillus spp., are recognized worldwide as a public health problem due to their potent carcinogenic, hepatotoxic, and immunosuppressive effect. Aflatoxin contamination in children is of particular concern given the negative impacts on health, development, and quality of life. The objective of this study was to validate the indirect competitive enzyme-linked immunosorbent assay (ic-ELISA) for the analysis of aflatoxin B1 (AFB1) in urine, as well as to evaluate the level of this mycotoxin in 150 urine samples from children living in Londrina, Paraná State, Brazil. The analytical performance parameters evaluated in the validation included specificity (matrix interference and comparison of curves with and without matrix), accuracy (recovery test), precision (repeatability and intermediate precision), robustness, sensitivity, linearity, limit of detection (LD) and limit of quantification (LQ). In the intralaboratory validation, matrix interference was analyzed with dilution factors of 2, 5 and 10 times, and the 5-fold dilution was selected to proceed. The standard curves prepared in the presence and absence of a matrix did not show significant differences in the percentage of linkage between the six points analyzed (p > 0.05). The average recovery rate for urine samples contaminated with AFB1 at concentrations of 1.0; 2.5 and 5.0 ng/mL were 100.00 ± 8.71%, 90.00 ± 2.00%, and 93.66 ± 1.52%, respectively. The coefficients of variation of the precision parameters were less than 15%. The regression equation obtained from seven standard curves was y=−14.89ln(x)+61.234y = -14.89 \ln(x) + 61.234y=−14.89ln(x)+61.234, with a coefficient of determination (R²) greater than 0.99. The LD was 0.096 ng/mL and the LQ was 0.115 ng/mL. AFB1 was detected in 39.3% of the samples analyzed, with concentrations ranging from 0.13 to 9.40 ng/mL and an average of 1.63 ± 1.57 ng/mL. Based on the results obtained, the validated ic-ELISA proved to be adequate for the determination of AFB1 in infant urine. Preliminary data suggest that the level of aflatoxin contamination in children in the region of Londrina, Paraná, Brazil, is low.

 

DOI:https://doi.org/10.56238/sevened2024.037-054


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This work is licensed under a Creative Commons Attribution-NonCommercial 4.0 International License.

Copyright (c) 2024 Angélica Tiemi Ishikawa, Estéfany Santos Redondo, Cassia Reika Takabayashi Yamashita, Elisabete Yurie Sataque Ono, Flávio Hiroshi Itano, Bianca Dorana de Oliveira Souza, Mario Augusto Ono, Osamu Kawamura, Elisa Yoko Hirooka, Eiko Nakagawa Itano

Author(s)

  • Angélica Tiemi Ishikawa
  • Estéfany Santos Redondo
  • Cassia Reika Takabayashi Yamashita
  • Elisabete Yurie Sataque Ono
  • Flávio Hiroshi Itano
  • Bianca Dorana de Oliveira Souza
  • Mario Augusto Ono
  • Osamu Kawamura
  • Elisa Yoko Hirooka
  • Eiko Nakagawa Itano