Salivary diagnostics is an emerging field that has progressed through several important developments in the past decade, including the publication of the human salivary proteome and the infusion of federal funds to integrate nanotechnologies and microfluidic engineering concepts into developing compact point-of-care devices for rapid analysis of this secretion. In this article, we discuss some of these developments and their relevance to the prognosis, diagnosis and management of periodontitis, as an oral target, and cardiovascular disease, as a systemic example for the potential of these biodiagnostics. Our findings suggest that several biomarkers are associated with distinct biological stages of these diseases and demonstrate promise as practical biomarkers in identifying and managing periodontal disease, and acute myocardial infarction. The majority of these studies have progressed through biomarker discovery, with the identified molecules requiring more robust clinical studies to enable substantive validation for disease diagnosis. It is predicted that with continued advances in this field the use of a combination of biomarkers in multiplex panels is likely to yield accurate screening tools for these diagnoses in the near future. Keywordsacute myocardial infarction; lab-on-a-chip; periodontitis; salivary diagnosis Overview of the field of salivary diagnosisThe analysis of blood and its components has been the mainstay for laboratory diagnostic procedures for several decades. However, other biological fluids are also utilized frequently for the diagnosis of disease, for example urine and cerebrospinal fluid, and thus, saliva could offer some distinct advantages in select situations [1][2][3][4][5][6]. Saliva is a hypotonic fluid NIH-PA Author ManuscriptNIH-PA Author Manuscript NIH-PA Author Manuscript composed mostly of water, electrolytes and organic molecules (i.e., amino acids, proteins and lipids). The water component is derived largely from the local capillary bed via intracellular diffusion, aquaporin water channels and extracellular routes [7,8]. Small neutral molecules from the serum enter by passive diffusion from the dense beds of capillaries surrounding and bathing the salivary glands. Electrolytes enter the saliva via osmotic gradients and are regulated by the rate of secretion, nature of the stimulus and level of mineralocorticoids in the circulation. The organic components of glandular saliva are derived largely from protein synthesis and are stored as granules within the acinar cells [4]. Because serum components of saliva are derived primarily from the local vasculature that originates from the carotid arteries [9], saliva has a prodigious fluid source that provides many, if not most, of the same molecules found in the systemic circulation. This makes saliva a potentially valuable fluid for the diagnosis of various systemic diseases (Figure 1).The recent cataloguing of the salivary proteome has availed considerable information that is potentially important for diagnostic applications ...

Aptamer biosensors have been immobilized on beads, introduced into micromachined chips on the electronic tongue sensor array, and used for the detection and quantitation of proteins. Aptamer chips could detect proteins in both capture and sandwich assay formats. Unlike most protein-based arrays, the aptamer chips could be stripped and reused multiple times. The aptamer chips proved to be useful for screening aptamers from in vitro selection experiments and for sensitively quantitating the biothreat agent ricin.

BackgroundMore than 35 million people in developing countries are living with HIV infection. An enormous global effort is now underway to bring antiretroviral treatment to at least 3 million of those infected. While drug prices have dropped considerably, the cost and technical complexity of laboratory tests essential for the management of HIV disease, such as CD4 cell counts, remain prohibitive. New, simple, and affordable methods for measuring CD4 cells that can be implemented in resource-scarce settings are urgently needed.Methods and FindingsHere we describe the development of a prototype for a simple, rapid, and affordable method for counting CD4 lymphocytes. Microliter volumes of blood without further sample preparation are stained with fluorescent antibodies, captured on a membrane within a miniaturized flow cell and imaged through microscope optics with the type of charge-coupled device developed for digital camera technology. An associated computer algorithm converts the raw digital image into absolute CD4 counts and CD4 percentages in real time. The accuracy of this prototype system was validated through testing in the United States and Botswana, and showed close agreement with standard flow cytometry (r = 0.95) over a range of absolute CD4 counts, and the ability to discriminate clinically relevant CD4 count thresholds with high sensitivity and specificity.ConclusionAdvances in the adaptation of new technologies to biomedical detection systems, such as the one described here, promise to make complex diagnostics for HIV and other infectious diseases a practical global reality.

In the last decade, saliva has been advocated as a non-invasive alternative to blood as a diagnostic fluid. However, use of saliva has been hindered by the inadequate sensitivity of current methods to detect the lower salivary concentrations of many constituents compared to serum. Furthermore, developments in the areas related to lab-on-a-chip systems for saliva-based point of care diagnostics are complicated by the high viscosity and heterogeneous properties associated with this diagnostic fluid. The biomarker C-reactive protein (CRP) is an acute phase reactant and a well-accepted indicator of inflammation. Numerous clinical studies have established elevated serum CRP as a strong, independent risk factor for the development of cardiovascular disease (CVD). CVD has also been associated with oral infections (i.e. periodontal diseases) and there is evidence that systemic CRP may be a link between the two. Clinical measurements of CRP in serum are currently performed with "high sensitivity" CRP (hsCRP) enzyme-linked immunosorbent assay (ELISA) tests that lack the sensitivity for the detection of this important biomarker in saliva. Because measurement of salivary CRP may represent a novel approach for diagnosing and monitoring chronic inflammatory disease, including CVD and periodontal diseases, the objective of this study was to apply an ultra-sensitive microchip assay system for the measurement of CRP in human saliva. Here, we describe this novel lab-on-a-chip system in its first application for the measurement of CRP in saliva and demonstrate its advantages over the traditional ELISA method. The increased sensitivity of the microchip system (10 pg ml(-1) of CRP with 1000-fold dilution of saliva sample) is attributed to its inherent increased signal to noise ratio, resulting from the higher bead surface area available for antigen/antibody interactions and the high stringency washes associated with this approach. Finally, the microchip assay system was utilized in this study to provide direct experimental evidence that chronic periodontal disease may be associated with higher levels of salivary CRP.

This paper provides the first direct structural evidence describing conjugated polymer self-assembly at the air-water interface. Grazing-incidence X-ray diffraction (GIXD) and X-ray reflectivity measurements on a number of derivatives of amphiphilic regioregular polythiophenes (e.g., poly(3′-dodecyl-3-(2,5,8trioxanonyl)-2′,5-bithiophene), polymer 1) show that these conjugated polymers self-assemble as 2-dimensional polycrystalline monolayers at the air/water interface with the amphiphilic polymers preorganized into rigid boards standing edge-on on the water surface. The monolayer consists of highly ordered (∼70% crystalline) domains, with a centered rectangular unit cell having the polymer backbone along the a-axis and the thiophene π-stack along the b axis with a distance of 3.85-3.94 Å depending on the applied surface pressure. These domains are connected by soft, more disordered boundaries. This is evidenced by the macroscopic compressibility of the entire LB film (C macro ≈ 4-7 m/N) being one order of magnitude larger than the microscopic compressibility (C micro ≈ 0.6 m/N) of the polycrystalline domains. The alkyl chains in the 3-position of the thiophene rings are in a crystallographically disordered state due to their cross-sectional mismatch with the packing of the thiophenes. The importance of having the side chains coupled in a regioregular fashion to the 3-position of the thiophene rings is evidenced by a dramatic increase in the coherence length of the crystalline domains for highly regioregular samples (>95% head-tail couplings) as compared to less regioregularly coupled polymers (∼80% head-tail couplings). Transfer to solid support by the Langmuir-Blodgett technique induces an overall orientation of the domains in the film, giving rise to a dichroic ratio of up to 4. Reflectionabsorption infrared spectroscopy (RAIRS) shows that the alkyl chains of transferred films are in an all-trans conformation with a locally ordered environment, having only few gauche defects.