Development and evaluation of a chemiluminescence immunoassay for type Ⅰ procollagen N-terminal peptide.

He Ren; Shuang Han; Xiang Chen; Shuaipeng LI; Zhonghu Bai

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Development and evaluation of a chemiluminescence immunoassay for type Ⅰ procollagen N-terminal peptide.

Author: He REN ¹ ; Shuang HAN ¹ ; Xiang CHEN ¹ ; Shuaipeng LI ¹ ; Zhonghu BAI ¹
Author Information

1. National Engineering Laboratory of Cereal Fermentation Technology, Jiangnan University, Wuxi 214122, Jiangsu, China.
Publication Type:Journal Article
Keywords: chemiluminescence; magnetic particle; performance evaluation; type Ⅰ procollagen N-terminal peptide
MeSH: Animals; Humans; Immunoassay; Luminescence; Mice; Mice, Inbred BALB C; Peptide Fragments/isolation & purification*; Procollagen/isolation & purification*
From: Chinese Journal of Biotechnology 2021;37(12):4406-4414
CountryChina
Language:Chinese
Abstract: To develop a magnetic nanoparticle chemiluminescence immunoassay (CLIA) for the determination of type Ⅰ procollagen N-terminal peptide (PINP) in human serum, we expressed a recombinant PINP-α1 protein in Corynebacterium glutamicum and used it as an immunogen to immunize BALB/c mice. We obtained three hybridoma cell lines that stably secret antibody against PINP-α1 protein. After further pairing and screening, we chose a monoclonal antibody 8C12 coupled with biotin as the capture antibody, and a monoclonal antibody 1F11 labeled horseradish peroxidase as the detection antibody. The antibodies combined with the serum samples, forming a sandwich complex which was used to detect the concentration of PINP in serum. After optimizing the conditions, we determined that the best working concentration of the capture antibody and the detection antibody were 3 μg/mL, and the incubation time was 30 minutes. The quantitative assay had a detection range of 5-1 100 ng/mL, with recovery rates between 93%-107% and the minimum detection limit of 1.22 ng/mL achieved. The intra-and inter-assay precisions were lower than 10%. The correlation coefficient of PINP results between this CLIA method and the Roche electrochemiluminescence immunoassay system was 0.906 2. Therefore, this CLIA method is specific and can be used to quantitatively detect the content of PINP in serum, which has the potential to become an auxiliary approach for bone disease examination.