Abstract
Over the past decades, paper-based lateral flow immunoassays (LFIAs) have been extensively developed for rapid, facile, and low-cost detection of a wide array of target analytes in a point-of-care manner. Conventional home pregnancy tests are the most significant example of LFAs, which detect elevated concentrations of human chorionic gonadotrophin (hCG) in body fluids to identify early pregnancy. In this work, we have upgraded these platforms to a higher version by developing a customized microfluidic paper-based analytical device (μPAD), as the new generation of paper-based point-of-care platforms, for colorimetric immunosensing. This will offer a cost-efficient and environmentally friendly alternative platform for paper-based immunosensing, eliminating the need for nitrocellulose (NC) membrane as the substrate material. The performance of the developed platform is demonstrated by detection of hCG (as a model case) in urine samples and subsequently indicating positive or negative pregnancy. A dual-functional silane-based composite was used to treat filter paper in order to enhance the colorimetric signal intensity in the detection zones of μPADs. In addition, microfluidic pathways were designed in a manner to provide the desired regulated fluid flow, generating sufficient incubation time (delays) at the designated detection zones, and consequently enhancing the obtained signal intensity. The presented approaches allow to overcome the existing limitations of μPADs in immunosensing and will broaden their applicability to a wider range of assays. Although, the application of the developed hCG μPAD assay is mainly in qualitative (i.e., positive or negative) detection of pregnancy, the semi-quantitative measurement of hCG was also investigated, indicating the viability of this assay for sensitive detection of the target hCG analyte within the related physiological range (i.e., 10–500 ng/mL) with a LOD value down to 10 ng/mL.
Highlights
Traditional lateral flow assays (LFIAs) are immunochromatographic paperbased platforms comprising a nitrocellulose (NC) membrane as a reaction surface with a high protein-binding capability, suitable for irreversible antibody immobilization
Conventional home pregnancy test LFAs are the most popular and widely commercialized tools for rapid, simple, and qualitative detection of elevated concentrations of human chorionic gonadotrophin in urine or serum samples, identifying pregnancy at early stages. hCG is a glycoprotein hormone normally generated by the placenta during pregnancy. hCG molecule is composed of 237 amino acids with a molecular mass of 36.7 kDa and two subunits, the alpha and beta
The normal serum hCG level is less than 10 ng/mL; the detection of hCG at this level is significant for early pregnancy identification
Summary
Traditional lateral flow (immuno) assays (LFIAs) are immunochromatographic paperbased platforms comprising a nitrocellulose (NC) membrane as a reaction surface with a high protein-binding capability, suitable for irreversible antibody immobilization. Conventional home pregnancy test LFAs are the most popular and widely commercialized tools for rapid, simple, and qualitative detection of elevated concentrations of human chorionic gonadotrophin (hCG) in urine or serum samples, identifying pregnancy at early stages. Secretion of hCG starts almost a week after fertilization, which reaches to its peak (1000 ng/mL) in 8 weeks of pregnancy. After this period, the hCG level begins to decline until it stabilizes. There are various methods available for detection of hCG (e.g., ELISA), the conventional LFA based tests offer numerous advantages, such as being rapid, simple, low cost, and easy to operate [3]
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