Single-Step FRET-Based Detection of Femtomoles DNA.

Sensitive detection of nucleic acids and identification of single nucleotide polymorphism (SNP) is crucial in diagnosis of genetic diseases. Many strategies have been developed for detection and analysis of DNA, including fluorescence, electrical, optical, and mechanical methods. Recent advances in fluorescence resonance energy transfer (FRET)-based sensing have provided a new avenue for sensitive and quantitative detection of various types of biomolecules in simple, rapid, and recyclable platforms. Here, we report single-step FRET-based DNA sensors designed to work via a toehold-mediated strand displacement (TMSD) process, leading to a distinct change in the FRET efficiency upon target binding. Using single-molecule FRET (smFRET), we show that these sensors can be regenerated in situ, and they allow detection of femtomoles DNA without the need for target amplification while still using a dramatically small sample size (fewer than three orders of magnitude compared to the typical sample size of bulk fluorescence). In addition, these single-molecule sensors exhibit a dynamic range of approximately two orders of magnitude. Using one of the sensors, we demonstrate that the single-base mismatch sequence can be discriminated from a fully matched DNA target, showing a high specificity of the method. These sensors with simple and recyclable design, sensitive detection of DNA, and the ability to discriminate single-base mismatch sequences may find applications in quantitative analysis of nucleic acid biomarkers.

Fatal factitious Cushing's syndrome and invasive aspergillosis: case report and review of literature.

OBJECTIVE: To present the first reported fatality from invasive aspergillosis related to factitious Cushing's syndrome. METHODS: We summarize the history, clinical findings, and outcome in a patient ultimately found to have factitious Cushing's syndrome. In addition, the dangers of fulminant infections in untreated Cushing's syndrome are analyzed relative to molecular and immunologic aspects, and the pertinent literature is reviewed. RESULTS: A 33-year-old female medical transcriptionist was admitted with rapidly fatal septic shock and diffuse pulmonary infiltrates. Autopsy revealed invasive pulmonary aspergillosis and atrophied adrenal cortices. On subsequent investigation, hidden bottles of prednisone were found throughout the patient's home. Factitious Cushing's syndrome has rarely been described and can be a difficult diagnosis to establish, but it is important to recognize this condition because of its potentially drastic consequences. Our understanding of the mechanism of immunosuppression from glucocorticoids related to the increased risk of invasive fungal infections is evolving. Factitious illness can manifest in numerous ways; therefore, health-care providers in all specialties should be familiar with epidemiologic, diagnostic, and treatment considerations for this illness. CONCLUSION: Endocrinologists should be aware of the possibility of factitious Cushing's syndrome because it can be an elusive and ultimately fatal condition.