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Updated: Jun 26, 2026
Immuno-fluorescence Assay of Leptospiral Surface-exposed Proteins
Published on: July 1, 2011
[Application of purified recombinant outer membrane lipoprotein LipL32 in detecting antibodies among leptospirosis
Guo-ying Xu1, Yan-sheng Yan, Zhi-shan Zhang
1Fujian Provincial Center for Disease Control and Prevention, Fuzhou 350001, China.
Objective:
To establish recombinant outer membrane lipoprotein LipL32-based antibody detection assays in identifying leptospirosis.
Methods:
Recombinant leptospiral outer membrane protein LipL32 was obtained by genetic engineering method. This purified protein was used in the indirect and sandwich ELISA assays to test the antibodies in sera of human beings and rats, and the results were compared with those obtained by microscopy agglutination test (MAT) and imported ELISA kit.
Results:
When the acute and convalescent phase specimens from 9 leptospiral patients were tested, the detected rates of three ELISAs were similar to the MAT. Among the 45 probable cases which MAT showed positive, 32 (71.11%) samples were positive by r32-I-ELISA, 36 (80.00%) by r32-S-ELISA, while 28.89% (13/45) samples were positive and 55.56% (25/45) were suspicious by D.A.I-ELISA. The specificity of r32-I-ELISA and r32-S-ELISA were 97.10% (67/69) for 69 specimens. 43 healthy specimens were negative by both r32-I-ELISA and r32-S-ELISA, 14 healthy specimens were negative by D.A.I-ELISA. Among 16 non-leptospirosis patients, two specimens were positive by r32-I-ELISA and r32-S-ELISA, D.A.I-ELISA and identified one positive specimen, while 12 specimens were suspicious by D.A.I-ELISA. For 10 syphilis specimens, data obtained through three ELISAs were in consistent with that by MAT. A sandwiched ELISA, using rLipL32 protein as the antigen was developed to detect rat sera. Considering MAT as standard test, the sensitivity and specificity were 86.75% (131/151), 99.19% (122/123) respectively with coincidence rate as 92.34% (253/274).
Conclusion:
The recombinant protein LipL32 had high immunoreactivity and could be used as an antigen for the detection of pathogenic leptospirosis. In summary, the novel sandwiched ELISA with rLipL32 showed similar sensitivity and specificity to that of MAT in the antibody detection of rat leptospirosis. It was suitable for large scales field sero-epidemiological studies.
Insights
Recombinant outer membrane lipoprotein LipL32 is effective for detecting leptospirosis antibodies. New sandwich ELISA assays using this protein show high sensitivity and specificity, suitable for large-scale epidemiological studies.
Area of Science:
- Immunology
- Infectious Diseases
- Biotechnology
Background:
- Leptospirosis is a significant zoonotic disease.
- Accurate serological diagnostic tools are crucial for leptospirosis surveillance.
- Outer membrane lipoproteins are key targets for antibody detection.
Purpose of the Study:
- To develop and validate recombinant LipL32-based antibody detection assays for leptospirosis.
- To evaluate the performance of indirect and sandwich ELISA methods using recombinant LipL32.
- To compare these novel assays with established diagnostic methods like MAT.
Main Methods:
- Genetic engineering was used to obtain recombinant leptospiral outer membrane protein LipL32.
- Indirect ELISA (r32-I-ELISA) and sandwich ELISA (r32-S-ELISA) were developed using purified rLipL32.
- Assays were tested on human and rat sera, comparing results with microscopy agglutination test (MAT) and an imported ELISA kit.
Main Results:
- Recombinant LipL32 ELISAs showed comparable detection rates to MAT in human acute and convalescent samples.
- r32-S-ELISA demonstrated high sensitivity (80%) and specificity (97.10%) in probable human leptospirosis cases.
- The sandwich ELISA for rat sera achieved 86.75% sensitivity and 99.19% specificity against MAT.
Conclusions:
- Recombinant LipL32 exhibits high immunoreactivity and is a suitable antigen for pathogenic leptospirosis detection.
- The novel sandwich ELISA using rLipL32 is a sensitive and specific tool for rat leptospirosis antibody detection.
- This assay is well-suited for large-scale field sero-epidemiological studies of leptospirosis.
