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Development and validation of enzyme-linked immunosorbent assay for anti-mouse pertussis immunoglobulin G using international reference anti-Bordetella pertussis mouse serum NIBSC 97/642.
Kang, Kyu-Ri; Kwon, Yi-Hyeon; Cho, Gyu-Won; Choi, Gi-Sub; Ji, Joon-Hwan; Kang, Hyun-Mi; Lee, Soo-Young; Kang, Jin-Han.
Afiliación
  • Kang KR; The Vaccine Bio Research Institute, Seoul St. Mary's Hospital, College of Medicine, The Catholic University of Korea, Seoul, Korea.
  • Kwon YH; The Vaccine Bio Research Institute, Seoul St. Mary's Hospital, College of Medicine, The Catholic University of Korea, Seoul, Korea.
  • Cho GW; The Vaccine Bio Research Institute, Seoul St. Mary's Hospital, College of Medicine, The Catholic University of Korea, Seoul, Korea.
  • Choi GS; CLIPS BnC, Research Center, Seoul, Korea.
  • Ji JH; CLIPS BnC, Research Center, Seoul, Korea.
  • Kang HM; The Vaccine Bio Research Institute, Seoul St. Mary's Hospital, College of Medicine, The Catholic University of Korea, Seoul, Korea.
  • Lee SY; Department of Pediatrics, College of Medicine, The Catholic University of Korea, Seoul, Korea.
  • Kang JH; The Vaccine Bio Research Institute, Seoul St. Mary's Hospital, College of Medicine, The Catholic University of Korea, Seoul, Korea.
Clin Exp Vaccine Res ; 13(3): 242-252, 2024 Jul.
Article en En | MEDLINE | ID: mdl-39144122
ABSTRACT

Purpose:

In this study, an in-house enzyme-linked immunosorbent assay (ELISA) was developed and validated. The titer of ELISA was calculated using the reference line (RFL) method based on the standard curve drawn using the international reference anti-mouse serum NIBSC (National Institute for Biological Standards and Control) 97/642. Materials and

Methods:

In the development step, signal to noise was depicted to select the buffers that showed the most appropriate ratio. In the validation step, standard range, precision, dilution linearity, and specificity were confirmed, and RFL and parallel line (PLL) methods were compared in precision and dilution linearity.

Results:

Coating concentration for plate was achieved at 0.1 µg/mL for pertussis toxin (PT), 0.15 µg/mL for filamentous hemagglutinin antigen (FHA), and 0.25 µg/mL for pertactin (PRN). The signal to noise ratio was 22.02 for PT, 14.93 for FHA, and 8.02 for PRN with 0.25% goat serum in phosphate-buffered saline (PBS) as a dilution buffer, and 2% skim milk in PBS as a blocking buffer. Based on the precision results, we assessed the lower limit of quantification by 1, 0.2, and 1.5 EU/mL concentration for PT, FHA, and PRN which met the ICH (International Council for Harmonization) M10 criteria of a 25% accuracy and total error of 40%. In specificity, homologous serum was spiked into heterologous serum and the accuracy met the criteria. There was no difference in the results between RFL and PLL calculations (p-value=0.3207 for PT, 0.7394 for FHA, 0.2109 for PRN).

Conclusion:

ELISA validated with RFL calculation method in this study is a relatively accurate assay for mouse humoral immunogenicity test.
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Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Idioma: En Revista: Clin Exp Vaccine Res Año: 2024 Tipo del documento: Article Pais de publicación:
Texto completo
Añadir a Mi BVS
Imprimir
XML
PubMed Links
Buscar en Google

Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Idioma: En Revista: Clin Exp Vaccine Res Año: 2024 Tipo del documento: Article Pais de publicación: