Native Hapten ELISA Kit for Brucellosis Diagnosis
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Solution Overview
Problem
Current serological tests for brucellosis in cattle often produce false positives due to cross-reactivity with other Gram-negative bacteria, and vaccination can interfere with diagnostic results, making it difficult to accurately differentiate between infected and vaccinated animals.
Innovation Solution
Development of an indirect ELISA kit based on the detection of antibodies against the Native Hapten antigen of Brucella, which is specific to infected animals and not produced by vaccination, thereby reducing false positives and improving diagnostic accuracy.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If traditional serological tests (Rose Bengal Test, Rivanol test, Milk Ring Test) are used for brucellosis detection, then the tests can detect antibodies against Brucella LPS, but they produce false positives due to cross-reactivity with other Gram-negative bacteria and vaccination interference
Solution Approach 1:
The patent extracts and uses only the Native Hapten (NH) antigen from the Brucella outer membrane, separating it from the LPS component. This extraction allows the test to detect antibodies specific to NH, which are produced during actual infection but not during vaccination, thereby eliminating false positives while maintaining diagnostic accuracy.
Solution Approach 2:
The patent focuses the diagnostic test on a specific local antigen (Native Hapten) within the Brucella structure rather than using general LPS antigens. This localized approach targets the specific immunological response caused by infection, distinguishing it from vaccination-induced responses, thus improving reliability and reducing false positives.
2Measurement precision
If LPS-based tests are used for brucellosis diagnosis, then the tests can identify antibodies against Brucella outer membrane components, but cross-reactivity with other Gram-negative bacteria reduces specificity
Solution Approach 1:
The patent extracts and isolates the Native Hapten antigen from the Brucella outer membrane, separating it from the LPS component. This extraction allows the test to detect antibodies specific to NH, which are produced during actual infection but not during vaccination, thereby eliminating false positives while maintaining diagnostic accuracy.
Solution Approach 2:
The patent focuses the diagnostic test on a specific local antigen (Native Hapten) within the Brucella structure rather than using general LPS antigens. This localized approach targets the specific immunological response caused by infection, distinguishing it from vaccination-induced responses, thus improving reliability and reducing false positives.
3Reliability
If vaccination with Brucella strains S19 or RB51 is performed, then the animals are protected against brucellosis, but the vaccination interferes with diagnostic test results
Solution Approach 1:
The patent extracts and uses only the Native Hapten (NH) antigen from the Brucella outer membrane, separating it from the LPS component. This extraction allows the test to detect antibodies specific to NH, which are produced during actual infection but not during vaccination, thereby eliminating false positives while maintaining diagnostic accuracy.
Solution Approach 2:
The patent focuses the diagnostic test on a specific local antigen (Native Hapten) within the Brucella structure rather than using general LPS antigens. This localized approach targets the specific immunological response caused by infection, distinguishing it from vaccination-induced responses, thus improving reliability and reducing false positives.
Applied Scientific Principles
This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.
Function Achieved in This Case
The kit effectively discriminates between truly positive animals and false positives, ensuring that only infected animals are identified, thereby preventing the spread of brucellosis and reducing economic losses in cattle breeding.
Implementation Method 1
The indirect enzyme-linked immunosorbent assay of anti-NH ELISA is a test that is fundamentally based on the determination of IgG isotype antibodies against the Brucella antigen named Native Hapten (NH) in samples of blood serum, whole milk and/or whey
Implementation Method 2
This kit includes microplates coated with the Native Hapten antigen in its raw form, the washing solutions, the sample diluent, the conjugate diluent, the substrate, the stop solution and the positive and negative controls, previously submitted to a lyophilization process
Data Source
AI summary
A diagnostic kit for a confirmatory assay using the indirect ELISA method that measures the levels of anti-Native Hapten antibodies produced during a real infection, thereby preventing large financial losses to livestock farm, by discerning “false positives” that present anti-LPS antibodies due to cross-reactions with enterobacteria and post-vaccinal antibodies for the diagnosis of bovine brucellosis in blood serum and individual milk (per animal) and bulk milk (tank), characterised by using the crude Native Hapten antigen, extracted from B. melitensis 16M strain, with no purification treatment and an effective adherence capacity, which is used to antigenize plates at a known concentration (1 μg per well), where using as reference positive and negative controls subjected to the lyophilisation (freeze drying) method to ensure their preservation, thereby avoiding the contamination and degradation of the antibodies present and ensuring the stability of the optical densities in said controls for a correct results interpretation of an indirect ELISA, are taken as reference. The lyophilisation method for controls that may be used in other diagnostic methods is also presented.