Establishment of a sandwich ELISA method for CHGA in saliva samples and its preliminary application in stress detection.
- Author:
Niqi SHAN
1
;
Shanshou LIU
2
;
Yuling WANG
1
;
Hui LIU
3
;
Shuai WANG
3
;
Yilin WU
4
;
Chujun DUAN
5
;
Hanyin FAN
4
;
Yangmengjie JING
6
;
Ran ZHUANG
7
;
Chunmei ZHANG
8
,
9
Author Information
1. Department of Immunology, Basic Medical Science Academy, Air Force Medical University, Xi'an 710032, China.
2. Department of Emergency, Xijing Hospital, Air Force Medical University, Xi'an 710032, China.
3. Department of General Surgery, Tangdu Hospital, Air Force Medical University, Xi'an 710038, China.
4. Department of Immunology, Basic Medical Science Academy, Air Force Medical University, Xi'an 710032; Medical School of Yan'an University, Yan'an 716000, China.
5. Department of Immunology, Basic Medical Science Academy, Air Force Medical University, Xi'an 710032; Institute of Medical Research, Northwestern Polytechnical University, Xi'an 710072, China.
6. Department of Immunology, Basic Medical Science Academy, Air Force Medical University, Xi'an 710032; College of Life Science, Yan'an University, Yan'an 716000, China.
7. Department of Immunology, Basic Medical Science Academy, Air Force Medical University, Xi'an 710032, China. *Corresponding authors, E-mail: fmmuzhr@fmmu.edu.cn.
8. Department of Immunology, Basic Medical Science Academy, Air Force Medical University, Xi'an 710032, China. *Corresponding authors, E-mail: hzcm1981@
9. com.
- Publication Type:Journal Article
- MeSH:
Saliva/metabolism*;
Enzyme-Linked Immunosorbent Assay/methods*;
Humans;
Animals;
Mice, Inbred BALB C;
Mice;
Chromogranin A/immunology*;
Antibodies, Monoclonal/immunology*;
Female;
Male;
Reproducibility of Results;
Adult
- From:
Chinese Journal of Cellular and Molecular Immunology
2025;41(4):324-330
- CountryChina
- Language:Chinese
-
Abstract:
Objective To establish a sandwich enzyme-linked immunosorbent assay (ELISA) method for the quantitative detection of Chromogranin A (CHGA) in saliva, and to explore its preliminary application in the testing of saliva samples. Methods Recombinant human CHGA protein was used to immunize BALB/c mice, and monoclonal antibodies (mAbs) were prepared and screened using conventional hybridoma technology. A double-antibody sandwich ELISA detection method was constructed, and the matrix effect of saliva samples was optimized. This method was then applied to detect the concentration of CHGA in the saliva of stressed individuals. Results Twenty-one stable hybridoma cell lines secreting high affinity anti-human CHGA antibodies were obtained. A pair of detection antibodies with the best effect was selected, and the optimal coating concentration was determined to be 10 μg/mL, with the optimal dilution of detection antibodies being 1:32 000. The accuracy and reproducibility of this method were verified, with both intra-batch and inter-batch variation coefficients less than 15×, and the recovery rate between 80× and 120×. The matrix effect was further optimized to make it suitable for saliva sample detection. Saliva samples from individuals in different stress states were collected, and the CHGA levels were detected using the method established in this study, indicating its potential to reflect the intensity of stress. Conclusion A reliable saliva CHGA ELISA detection method has been successfully established, and its potential as a biomarker in stress-related research has been preliminarily explored.
