Development of a paper-based lateral flow prothrombin assay.

Disorders of haemostasis result in both excessive bleeding and clotting and are a major global cause of morbidity and mortality, particularly in the developing world. A small number of simple tests can be used to screen and monitor for such dysfunctions, one of which is the prothrombin time (PT) test and associated International Normalisation Ratio (INR). PT/INR is routine in hospital laboratories in developed countries, and can also be performed using point-of-care instruments. However, neither of these approaches is appropriate in low-resource settings. Significant interest has grown in paper-based devices to form the basis of simple and low-cost assays that may have the potential for application in such environments. This study describes the development of a simple, low-cost, paper-based lateral flow prothrombin assay. The assay employed wax printing on chromatography paper to define test channels, with deposition of thromboplastin reagent and calcium chloride onto the resulting strips. These were placed in a test housing and measurement of the flow rates of deposited plasma samples were performed in triplicate. The flow dynamics of the assay was optimised according to the type of paper substrate used, the nature and quantity of the thromboplastin reagent, the amount of calcium chloride required, and the volume of sample employed. An optimised assay configuration demonstrated a dynamic range of 6 mm between normal and factor-deficient plasmas. The assay showed good correlation with laboratory-based PT assay (Yumizen G200) in artificial plasmas in the 9.8 to 36 s range (r2 = 0.8112). The assay also demonstrated good dynamic range and correlation in patient plasma samples in comparison with hospital PT, with a range of 9.8 to 45 s (r2 = 0.7209).

Paper-Based Lateral Flow Device for the Sustainable Measurement of Human Plasma Fibrinogen in Low-Resource Settings.

Fibrinogen concentration is a major determinant of both clotting and bleeding risk. Clotting and bleeding disorders cause extensive morbidity and mortality, particularly in resource-poor and emergency settings. This is exacerbated by a lack of timely intervention informed by measurement of fibrinogen levels under conditions such as thrombosis or postpartum haemorrhage. There is an absence of simple, rapid, low-cost, and sustainable diagnostic devices for fibrinogen measurement that can be deployed in such environments. Paper-based analytical devices are of significant interest due to their potential for low-cost production, ease of use, and environmental sustainability. In this work, a device for measuring blood plasma fibrinogen using chromatography paper was developed. Wax printing was used to create hydrophobic structures to define the test channel and sample application zone. Test strips were modified with bovine thrombin. Plasma samples (22 µL) were applied, and the flow rate was monitored over 5 min. As the sample traversed the strip, clotting was induced by the conversion of soluble fibrinogen to insoluble fibrin. The flow rate and distance travelled by the sample were dependent on fibrinogen concentration. The device was able to measure fibrinogen concentration in the range of 0.5-7.0 ± 0.3 mg/mL (p < 0.05, n = 24) and had excellent correlation with laboratory coagulometry in artificial samples (r2 = 0.9582, n = 60). Devices were also stable at 4-6 °C for up to 3 weeks.