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We have developed a new method for unprocessed biological specimens as templates directly into the TaqMan assay. Saliva was needed to be put on a water-soluble paper and dried, because foreign substances, such as a filter paper, hinder fluorescence detection through the assay. Genotyping of alcohol metabolism-related genes ADH1B (rs1229984) and ALDH2 (rs671) polymorphisms was, subsequently, performed by TaqMan PCR assay using dried saliva in the present investigation. The optimized technique was tested on 114 samples of alcoholic patients. The PCR-RFLP methods with purified DNA from blood samples were employed for validation of the assay. Upon validation, complete concordance was observed between the two independent results. These results highlight the ability of TaqMan PCR assays using dried saliva on water-soluble paper in genotyping of ADH1B and ALDH2 genes. Our results showed a rapid, simple, reliable, and cost-effective method for SNP genotyping of mutations in ADH1B and ALDH2 genes. This will be very useful for large-scale association studies in various fields. [Original].

We have developed a new method for directly using unprocessed biological specimens as templates for the TaqMan assay. DNA extraction and purification had been believed to be required for the assay, but our new method could avoid hindering fluorescence detection, even if the templates were used directly. Saliva was needed to be put on water-soluble paper and dried, and hairs were cut to be about 10 mm long. This method could reduce both the time and effort involved, and also the risk of contamination. It should prove to be very valuable for genetic diagnoses in various fields.

Recent advances in electrochemical analysis on filter paper exemplify the versatility of this substrate for high performance testing. Its low-cost, light-weight, and environmentally friendly properties make it particularly attractive for applications in addressing health and environmental safety needs in low-resource settings and developing countries. However, the main drawback to sensitive electrochemical testing is the use of a potentiostat, a bench-top instrument that is extremely expensive, thereby negating the some of the benefits of paper-based devices. Hence there is a need to develop paper-devices for use with handheld, portable device readers that can extract quantitative readouts. In this study, we developed a method to use micro-paper electrochemical devices, or µPEDs, with a glucose meter, which are used for personal monitoring of blood glucose levels. Ethanol was chosen as a model target analyte due to its importance in the global issue of road safety. µPEDs were simple in design and could be tested with a potentiostat. We observed that inclusion of the stabilizer trehalose was critical to preparing µPEDs for later analysis. In addition, an NAD+-dependent enzyme was used to impart selectivity to the biosensor, which also represents a class of enzymes with targets relevant to the health and food industry.

This paper describes the use of the electrostatic layer-by-layer (LbL) technique for the preparation of bioanodes with potential application in ethanol/O2 biofuel cells. More specifically, the LbL technique was employed for immobilization of dehydrogenase enzymes and polyamidoamine (PAMAM) dendrimers onto carbon paper support. Both mono (anchoring only the enzyme alcohol dehydrogenase, ADH) and bi-enzymatic (anchoring both ADH and aldehyde dehydrogenase, AldDH) systems were tested. The amount of ADH deposited onto the Toray® paper was 95 ng cm−2 per bilayer. Kinetic studies revealed that the LbL technique enables better control of enzyme disposition on the bioanode, as compared with the results obtained with the bioanodes prepared by the passive adsorption technique. The power density values achieved for the mono-enzymatic system as a function of the enzyme load ranged from 0.02 to 0.063 mW cm−2 for the bioanode containing 36 ADH bilayers. The bioanodes containing a gas diffusion layer (GDL) displayed enhanced performance, but their mechanical stability must be improved. The bi-enzymatic system generated a power density of 0.12 mW cm−2. In conclusion, the LbL technique is a very attractive approach for enzyme immobilization onto carbon platform, since it enables strict control of enzyme disposition on the bioanode surface with very low enzyme consumption.

Superabsorbent polymers are shown to be an excellent medium for the detection of enzymes used in enzyme assays. They offer increased sensitivity of 2-10 times over conventional blotter medium. Their usefulness is demonstrated with a quick, inexpensive, visual test for alcohol in urine with sensitivity approaching 0.001% (w/v). Alcohol levels in 500 random urine samples were determined and the levels ranged from 0.06 to less than 0.001%, with the majority of positives being near 0.005%.