Immunoassays for differential detection of clostridium difficile
Inventors
Pollock, Nira • Kelly, Ciaran • Duffy, David C. • Song, Linan • Zhao, Mingwei
Assignees
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Publication Number
US-11434264-B2
Publication Date
2022-09-06
Expiration Date
Abstract
Ultrasensitive and quantitative assays for detecting toxins of Clostridium difficile, which may involve digital ELISA, are provided. Also provided herein are differential detection assays that allow for distinguishing toxin B of highly virulent Clostridium difficile strains from toxin B of less virulent strains.
Core Innovation
The invention provides methods for measuring multiple toxins of Clostridium difficile in a stool sample suspected of containing a C. difficile strain and/or a toxin thereof. The methods measure a level of toxin B in the stool sample using an optical interrogation-based ELISA with a limit of detection of no greater than 20 pg/mL for toxin B in stool, and measure a level of toxin A in the stool sample using an optical interrogation-based ELISA with a limit of detection of no greater than 10 pg/mL for toxin A in stool.
The described optical interrogation-based ELISA uses affinity capture objects and an optical interrogation in which toxin associates with a statistically significant fraction of the plurality of capture objects while a statistically significant fraction remains unassociated. The optical interrogation provides a measure indicative of how many capture objects are associated with toxin, and a level of toxin is determined based on this measure for toxin B and for toxin A.
The invention further addresses C. difficile infection management by selecting or determining subjects for treatment based on symptom-associated diagnosis and quantitative stool toxin testing. Diagnosis includes determining that a subject manifests a symptom associated with C. difficile infection and determining a level of toxin A, toxin B, or both in a stool sample using the optical interrogation-based assay with specified limits of detection, and in one treatment workflow the diagnosis is confirmed based on the test results and a suitable anti-C. difficile drug is administered.
Claims Coverage
The document includes four independent claims, each centered on optical interrogation-based ELISA measurement of toxin A and/or toxin B in stool and quantitative limits of detection that gate diagnosis and treatment. Across the independent claims, the inventive features include dual-toxin measurement in stool with specified limits of detection, optical interrogation-based assay using capture-object association and determination from associated-object measures, and using the quantified toxin results to select or confirm infection for administering a suitable anti-C. difficile drug.
Dual-toxin optical interrogation-based ELISA in stool with specified limits of detection
A method for measuring a level of multiple toxins of Clostridium difficile in a stool sample suspected of containing a C. difficile strain and/or a toxin thereof, comprising providing the stool sample; measuring a level of toxin B using an optical interrogation-based ELISA having a limit of detection of no greater than 20 pg/mL for toxin B in stool; and measuring a level of toxin A using an optical interrogation-based ELISA having a limit of detection of no greater than 10 pg/mL for toxin A in stool.
Capture-object association quantified by optical interrogation for toxin levels
A method for determining levels of toxin A and/or toxin B in a stool sample by exposing capture objects with binding surfaces to toxin A/B; associating toxin A/B with some capture objects; using optical interrogation to determine a number of associated capture objects; and calculating toxin levels from that measure.
Treating C. difficile infection based on symptom diagnosis and quantitative optical assay thresholds for toxin A/toxin B
A method for treating Clostridium difficile infection in a subject, comprising selecting a subject having a diagnosis of C. difficile infection, the diagnosis being made by determining that the subject manifests a symptom associated with C. difficile infection and determining a level of toxin A, toxin B, or both in a stool sample using an optical interrogation-based assay having a limit of detection of about no greater than 10 pg/mL for toxin A in stool and no greater than 20 pg/mL for toxin B in stool; and administering a suitable anti-C. difficile drug to the subject.
Treating based on confirmed infection determined from quantified toxin levels using optical interrogation-based assay
A method for treating Clostridium difficile infection in a subject, comprising determining that the subject manifests a symptom associated with C. difficile infection; determining a level of toxin A, toxin B, or both in a stool sample using an optical interrogation-based assay having a limit of detection of about no greater than 10 pg/mL for toxin A in stool and no greater than 20 pg/mL for toxin B in stool; determining based on the results of step (ii) that the subject has C. difficile infection; and administering a suitable anti-C. difficile drug to the subject.
Overall claim coverage is focused on optical interrogation-based ELISA measurement of toxin A and toxin B in stool with specified limits of detection, implemented through capture-object association quantified by optical interrogation. Treatment claim coverage uses symptom-associated diagnosis and the quantitative toxin assay results to define eligibility or confirmation for administering a suitable anti-C. difficile drug.
Stated Advantages
Enables measuring levels of toxin B and toxin A in stool using an optical interrogation-based ELISA with specified limits of detection (no greater than 20 pg/mL for toxin B and no greater than 10 pg/mL for toxin A).
Supports determining infection status based on quantitative toxin A/toxin B results from the optical interrogation-based assay.
Enables administering a suitable anti-C. difficile drug to subjects based on diagnosis supported by symptom-associated determination and quantitative stool toxin testing.
Documented Applications
Clinical diagnosis and treatment selection/confirmation for C. difficile infection in a subject using symptom-associated diagnosis plus quantitative optical interrogation-based stool toxin measurement for toxin A and/or toxin B.
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