Journal of Shandong University (Health Sciences) ›› 2018, Vol. 56 ›› Issue (10): 51-57.doi: 10.6040/j.issn.1671-7554.0.2018.1027

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Construction and screening of CA50 targeted antibody library and establishment of immunoassay system

DU Lutao1,2, ZHAO Qiaohui3, TIAN Xiaoping3, WU Xuewei4, FU Guangyu4, QU Hai4, MIAO Yongjun4, WANG Lili5, ZHENG Guixi5, GUO Haiyang6, LI Guilin3, WANG Chuanxin1,2   

  1. 1. Department of Clinical Laboratory, The Second Hospital of Shandong University, Jinan 250033, Shandong, China;
    2. Tumor Marker Detection Engineering Laboratory of Shandong Province, Jinan 250033, Shandong, China;
    3. Zhengzhou Immuno Bio-Tech Co., Ltd, Zhengzhou 450016, Henan, China;
    4. Autobio Diagnostics Co., Ltd, Zhengzhou 450016, Henan, China;
    5. Department of Clinical Laboratory, Qilu Hospital of Shandong University, Jinan 250012, Shandong, China;
    6. Princess Margaret Cancer Centre/University Health Network, Toronto, M5G 1L7, Ontario, Canada
  • Received:2018-09-04 Published:2022-09-27

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Abstract: Objective To obtain antibodies against CA50 antigen with high affinity and specificity, and to establish a diagnostic kit so as to enhance the specificity and sensitivity of the diagnosis of tumor related diseases. Methods The mouse single chain antibody(ScFv)genes were obtained by PCR and cloned into pcomb3XSS phagemids. ScFv phage antibody library was obtained by electrotransformation. CA50 antigen containing epitope LST a(sialic acid lactose-N-four sugar a)was used for the elutriation screening. Random clone sequencing results showed that the antibody library 山 东 大 学 学 报 (医 学 版)56卷10期 -杜鲁涛,等. CA50靶向抗体文库的构建筛选及应用评价 \=-had good diversity. The human mouse chimeric antibody was constructed by overlap-PCR. The double antibody sandwich method CA50 kit was established after antibody expression purification and matching screening. Meanwhile, 54 clinical positive samples and 146 negative control samples were collected. The CA50 kit established was used for comparation with Roches CA50 kit to evaluate the detection correlation. Results The phage antibody library had a capacity of 2.5×108, and had a good diversity. The positive clones of 24 strains were identified by PCR. The sequencing results showed that 20 strains were correct and the sequences were different. After 3 rounds of panning, 5 positive clones were screened out, and the correlation between the results of CA50 kit and Roches CA50 kit was 0.98. Conclusion The antibodies with high specificity and affinity against CA50 were successfully screened by phage library technology, which made the application of CA50 possible in tumor diagnosis.

Key words: CA50 targets, Diagnosis, Antibody library, Evaluation

CLC Number: 

  • R392-33
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