Direct detection of genetic biomarkers in tissue and body fluids without complex target extraction and amplification processes can revolutionize nucleic acid-based diagnostics by enabling the use of this technology at the point-of-care. The development of point-of-care diagnostics is important to increase access to early treatment in underserved populations in low to middle income countries, which are disproportionally affected by infectious diseases and increasingly affected by certain types of cancer. The main obstacle to the development of such technologies is the low concentration of target sequences that makes this goal challenging. We report a method for direct detection of pathogen RNA in blood lysate using a bioassay using surface-enhanced Raman spectroscopy (SERS)-based detection assay that can be integrated in a “lab-in-a-stick” portable device. We could directly detect synthetic target with a limit of detection of 200 fM and, more importantly, we detected P. falciparum malaria parasite RNA directly in infected red blood cells lysate. Additionally, this paper will discuss the use of the developed assay for the identification of head and neck squamous cell carcinoma (HNSCC), which is an increasingly prevalent malignancy in low to middle income countries.
Gregory Palmer, Hengtao Zhang, Chen-Ting Lee, Husam Mikati, Joseph Herbert, Marlee Krieger, Jesko von Windheim, Dave Koester, Daniel Stevenson, Daniel Rocke, Ramon Esclamado, Alaatin Erkanli, Nirmala Ramanujam, Mark Dewhirst, Walter Lee
Diffuse reflectance spectroscopy (DRS) represents a quantitative, noninvasive, nondestructive means of assessing vascular oxygenation, vascularity, and structural properties. However, it is known that such measurements can be influenced by the effects of pressure, which is a major concern for reproducible and operator-independent assessment of tissues. Second, regular calibration is a necessary component of quantitative DRS to account for factors such as lamp decay and fiber bending. Without a means of reliably controlling for these factors, the accuracy of any such assessments will be reduced, and potentially biased. To address these issues, a self-calibrating, pressure-controlled DRS system is described and applied to both a patient-derived xenograft glioma model, as well as a set of healthy volunteers for assessments of oral mucosal tissues. It was shown that pressure had a significant effect on the derived optical parameters, and that the effects on the optical parameters were magnified with increasing time and pressure levels. These findings indicate that not only is it critical to integrate a pressure sensor into a DRS device, but that it is also important to do so in an automated way to trigger a measurement as soon as possible after probe contact is made to minimize the perturbation to the tissue site.
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