Development of an Indirect ELISA Kit for Measurement of Circulating Thyroid Peroxidase Levels in Patients with Hypothyroidism

Authors

• Aulanni’am Aulanni’am, Department of Chemistry, Faculty of Science Technology and Mathematics, Brawijaya University, Malang 65145, IndonesiaFollow
• Devi Ramadani, Department of Chemistry, Faculty of Science Technology and Mathematics, Brawijaya University, Malang 65145, IndonesiaFollow
• Almas Dwi Khairana, Department of Chemistry, Faculty of Science Technology and Mathematics, Brawijaya University, Malang 65145, IndonesiaFollow
• Saidun Fiddaroini, Department of Chemistry, Faculty of Science Technology and Mathematics, Brawijaya University, Malang 65145, IndonesiaFollow
• Muhammad Fikri Nur, Department of Chemistry, Faculty of Science Technology and Mathematics, Brawijaya University, Malang 65145, IndonesiaFollow
• Andreas Budi Wijaya, Faculty of Medicine, Malang State University, Malang 65145, Indonesia
• Rulli Rosandi, Faculty of Medicine, Brawijaya University, Malang 65145, Indonesia
• Dyah Kinasih Wuragil, Faculty of Veterinary Medicine, Brawijaya University, Malang 65151, Indonesia
• Wibi Riawan, Faculty of Medicine, Brawijaya University, Malang 65145, Indonesia

Abstract

Hypothyroidism, characterized by insufficient thyroid hormone production, affects approximately 5% of the population, especially women, thus increasing pregnancy-related risks. Conventional screening mainly measures thyroid-stimulating hormone (TSH) and free thyroxine (FT4). Detecting the presence of anti-thyroid peroxidase (anti-TPO) antibodies indi-cates autoimmune thyroid disease. This study developed an indirect enzyme-linked immunosorbent assay (ELISA) kit for detecting circulating TPO using polyclonal antibodies generated in rabbits. TPO was isolated from positive sera of pa-tients with hypothyroidism, purified by electroelution, and confirmed as a 52-kDa protein by Western blot. Rabbits immunized with this protein produced polyclonal antibodies with the highest titer (OD = 0.334) at week 8. Dot blot analysis demonstrated a detection capability up to a serum dilution of 1:80. The developed indirect ELISA showed reliable sensi-tivity and indicates potential cost-effectiveness. These findings demonstrate its potential as a diagnostic tool for early hypothyroidism detection and monitoring, providing a foundation for further clinical validation and broader application.

Recommended Citation

Aulanni’am, Aulanni’am; Ramadani, Devi; Khairana, Almas Dwi; Fiddaroini, Saidun; Nur, Muhammad Fikri; Wijaya, Andreas Budi; Rosandi, Rulli; Wuragil, Dyah Kinasih; and Riawan, Wibi (2026) "Development of an Indirect ELISA Kit for Measurement of Circulating Thyroid Peroxidase Levels in Patients with Hypothyroidism," Makara Journal of Science: Vol. 30: Iss. 3, Article 5.
DOI: 10.7454/mss.v30i3.2728
Available at: https://scholarhub.ui.ac.id/science/vol30/iss3/5