Methods and materials for the detection of Leishmania infection

Inventors

Houghton, Raymond L. • Reed, Steven G. • Raychaudhuri, Syamal

Assignees

InBios International Inc

Abstract

The present invention provides rapid diagnostic assays for the detection of Leishmania which can readily be used in the field, leading to more rapid treatment. In certain embodiments, the inventive assays, including ELISA and lateral flow assays, employ antibodies that may be effectively employed to detect the Leishmania major antigen TSA, which is present in both promastigotes grown in culture and in amastigotes. Such assays may be employed to detect the presence of cutaneous leishmaniasis in a subject using scrapings, biopsies, and/or aspirates taken from cutaneous lesions.

Core Innovation

The invention concerns rapid field-deployable immunoassays for detecting cutaneous leishmaniasis using antibodies that bind Leishmania major polypeptides associated with amastigotes. In particular, the immunoassays use antibodies specific for a polypeptide identified as SEQ ID NO: 1 (TSA/thiol specific antigen/peroxidoxin) and provide antibody-based detection formats including lateral flow assays and ELISA-based sandwich detection approaches.

A central feature is a sandwich immunodetection concept in which a labeled polyclonal antibody specific for a polypeptide of SEQ ID NO: 1 is used to form an antibody-polypeptide conjugate, and the conjugate is captured by a monoclonal antibody specific for SEQ ID NO: 1. The monoclonal antibody is raised against L. major amastigotes, and the antigen recognition is described as involving a conformational epitope associated with amastigotes.

The disclosed assay system also includes additional immunoassay formats and alternative antibody targets. It describes alternative assays using antibody WIC79.3 against L. major lipophosphoglycan (LPG) to detect secreted proteins/promastigotes (not amastigotes) and further describes kit components such as lateral flow/dipstick configurations and assay components for immunoassay operation. Supporting examples describe reactivity and specificity of TSA- and WIC79.3-based assays for detecting relevant Leishmania targets in the presence of non-target controls.

Claims Coverage

Two independent methods are presented, both covering sandwich immunodetection of cutaneous leishmaniasis using antibodies specific for a polypeptide of SEQ ID NO: 1. Across the independent claims, the inventive features primarily concern the orientation of labeled antibody (polyclonal versus monoclonal), capture antibody source (monoclonal raised against L. major amastigotes versus polyclonal), and detection of the resulting antibody-polypeptide conjugate.

The claim set covers sandwich immunodetection of cutaneous leishmaniasis centered on a polypeptide of SEQ ID NO: 1, with the core difference between the two independent claims being whether the labeled antibody is the polyclonal antibody or the monoclonal antibody, while maintaining capture by the complementary antibody and using a monoclonal antibody raised against L. major amastigotes.

Stated Advantages

Documented Applications