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Diagnosis of leptospirosis by recombinant antigen based single serum dilution ELISA
Sohini Dey1, C Madhan Mohan, P Ramadass
1Division of Animal Biotechnology, Indian Veterinary Research Institute, Bareilly, India. sohinimadhan@yahoo.com
Background & Objectives:
Leptospirosis, a zoonosis with a worldwide distribution is an acute febrile illness caused by spirochaetes of the pathogenic Leptospira interrogans. Microscopic agglutination test (MAT), the reference method for diagnosis was successively done to evaluate the modified ELISA which was developed with the recombinant LipL32 antigen for the detection of anti-leptospiral antibodies in human serum samples.
Methods:
The recombinant LipL32 antigen was developed from the serovar Pomona strain Pomona of the pathogenic L. interrogans species. The predicted titre at a single working dilution was plotted against the observed antiserum titre. Subsequently, predicted antibody activity titres were determined directly from the standard curve by solving the regression line equation. The relative sensitivity, specificity and accuracy of the single dilution ELISA for the detection of anti-leptospiral antibodies were determined in comparison to the MAT.
Results:
A linear relationship was found between the predicted antibody titres at a single working dilution of 1:250 and the corresponding observed serum titres by the standard serial-dilution method. Regression analysis was used to determine a standard curve from which an equation was derived that allowed demonstration of the mentioned correlation. The equation was then used to convert the corrected absorbance readings of the single working dilution directly into the predicted ELISA antibody titres. A high level of sensitivity of 96 per cent and specificity of 91 per cent between ELISA and MAT titres was found. The kappa value was almost 1.0 indicating perfect agreement.
Interpretation & Conclusions:
The r LipL32 ELISA was proved to be sensitive, specific and accurate as compared to the standard MAT and the test could be efficiently utilized as a screening test for a large number of human serum samples for the detection of leptospiral antibodies.
Insights
A modified ELISA using recombinant LipL32 antigen accurately detects leptospirosis antibodies in human serum. This sensitive and specific test, comparable to the Microscopic Agglutination Test (MAT), is suitable for large-scale screening.
Area of Science:
- Medical Microbiology
- Immunodiagnostics
- Zoonotic Diseases
Background:
- Leptospirosis is a globally distributed zoonotic disease caused by pathogenic *Leptospira interrogans*.
- The Microscopic Agglutination Test (MAT) is the gold standard for leptospirosis diagnosis.
- There is a need for efficient diagnostic methods for widespread screening.
Purpose of the Study:
- To evaluate a modified Enzyme-Linked Immunosorbent Assay (ELISA) for detecting anti-leptospiral antibodies in human serum.
- To assess the diagnostic performance of the modified ELISA using recombinant LipL32 antigen compared to MAT.
Main Methods:
- Recombinant LipL32 antigen was developed from *L. interrogans* serovar Pomona.
- A single working dilution ELISA was optimized and validated against serial dilution MAT.
- Regression analysis was used to establish a standard curve for predicting antibody titers.
Main Results:
- A linear correlation was observed between predicted and observed antibody titers using the single dilution ELISA.
- The modified ELISA demonstrated high sensitivity (96%) and specificity (91%) compared to MAT.
- A kappa value close to 1.0 indicated perfect agreement between the ELISA and MAT.
Conclusions:
- The recombinant LipL32 ELISA is a sensitive, specific, and accurate diagnostic tool for leptospirosis.
- This ELISA method is efficient for screening large numbers of human serum samples for leptospiral antibodies.
- The developed ELISA offers a reliable alternative for leptospirosis diagnosis.
