Project Summary/Abstract
HIV infection directly impacts both Mycobacterium tuberculosis pathogenesis and disease severity. Timely and
accurate diagnosis of active tuberculosis (TB) disease is critical for people living with HIV (PLWH) as TB
disease accounts for one third of all HIV-related deaths. Yet, currently there are no diagnostic solutions for
active TB disease that are rapid, inexpensive, and highly sensitive; and solutions that are available are
designed primarily as sputum-based solutions and optimized to detect pulmonary disease. This exacerbates
the diagnostic gap for PLWH, as PLWH are more likely to present with paucibacillary and extrapulmonary
forms of the disease. Novel point-of-care, rapid and accurate diagnostic solutions that have been optimized
and tested among PLWH are critically needed to reduce treatment delays and improve treatment outcomes.
Our central hypothesis is that the identification of the M. tuberculosis specific antigen, 10-kDa culture filtrate
protein or CFP-10, in circulating body fluids, specifically blood, is indicative of actively replicating bacteria, and
the detection of CFP-10 could potentially be used to diagnose both pulmonary and extrapulmonary disease.
Historically direct detection of M. tuberculosis specific antigens has been hampered by the expense and
complexity of detecting ultra-low concentrations of circulating antigens in biofluids. However, recent
technological advancements in chip-based electrochemical immunosensors have resulted in increased
sensitivity, options for miniaturization, and improved operational simplicity allowing for the detection of ultra-low
concentrations of target antigens in micro-volume clinical specimens.
Using seed funding from SD-CFAR our team developed a prototype Mycobacteria tuberculosis antigen
detection assay that successfully detected the M. tuberculosis specific antigen CFP-10 in a limited number of
clinical samples from patients with culture confirmed TB. While the initial success of this prototype assay was
very promising, more data is needed prior to any further clinical validation studies. In this proposed study we
aim to 1) determine the analytical performance of the prototype assay by conducting a series of laboratory-
based experiments using contrived samples to standardize the assay, assess its sensitivity and performance
parameters (dynamic range, precision, and repeatability), determine the specificity of the assay in the presence
of non-tuberculosis mycobacterium, and evaluate its storage stability: and 2) we will conduct a limited clinical
performance study of the prototype assay using 144 previously collected (72 from PLWH) and well
characterized clinical samples from the R2D2 network (with a pre-selected TB positivity of 50%). The low cost
and relatively simple approach of the prototype assay make it ideal for use at the clinic level as a low-tech,
point-of-care diagnostic. If ultimately successful, this assay has the potential to fundamentally change global
TB diagnostics.