Multiplex lateral flow assays are one of the greatest advancements in the world of rapid diagnostics, achieving the performance of several tests in one. These tests meet the basic requirements of increasing ease of use, low detection limit, and high specificity, as they combine the use of novel strategies, such as the exploitation of multiple detection labels, and a variety of amplification methods. These tests have proven their usefulness in many different areas, including clinical diagnostics, food, and environmental monitoring. In this review paper, we attempt to highlight and discuss the predominant changes in multianalyte LFAs, as related to their principle, their development, and their combination with other methods. Attention is paid to their flexibility and the challenges associated with the use of LFA arrays, including strategies to improve the detectability, sensitivity, and reliability of the assays. Therefore, this review emphasizes the current advances in the field to underline the possible impact of multiplex LFAs on the future of diagnostics and analytical sciences.

Liquid biopsy is an efficient diagnostic/prognostic tool for tumor-derived component detection in peripheral circulation and other body fluids. The rapid assessment of liquid biopsy techniques facilitates early cancer diagnosis and prognosis. Early and precise detection of tumor biomarkers provides crucial information about the tumor that guides clinicians towards effective personalized medicine. Point-of-care-testing remains still a great challenge in cancer diagnostics. Liquid biopsy is a promising alternative to tissue biopsy with the great advantages of less invasion and real-time monitoring of the disease, also providing information about tumor heterogeneity. The field is continuously and rapidly expanding. Numerous sophisticated biosensors have been developed targeting several biomarkers to achieve low detection limits, increased specificity and robustness. Current biosensors include mainly optical sensors, such as colorimetric, fluorescent, SPR, SERS and lateral flow assays. Electrochemical sensors have also been developed, providing very low detection limits. Colorimetric sensors exhibited simplicity in signal interpretation, while fluorescent sensors contributed to low analysis times, and SPR/SERS enabled label-free and rapid analysis. Novel target amplification and signal enhancement techniques have been exploited to increase the detectability of the sensors. In this context, this review is focused on the recent advances in biosensing technology for cutting-edge liquid biopsy applications towards point-of-care testing.

Triple-negative breast cancer (TNBC) is a highly aggressive type of cancer with limited therapeutic options. However, this type of cancer cells has shown overexpression of folate receptors, which bind with folic acid (FA) with high affinity. This feature can be used for targeting of nanocarriers, such as liposomes. In order to further examine the potential of increased efficacy by targeting the folate receptor, we prepared folate conjugated liposomes (DSPC/Chol/PEG/DSPE-PEG-FA) and loaded them with doxorubicin (DOX), an anticancer drug. For this, we first synthesized and verified the conjugate between FA and PEG-lipid (FA-PEG-lipid). After that, liposomes were prepared with thin film hydration method followed by probe sonication. Three different types of targeted liposomes having different concentrations of FA-PEG-lipid in their membranes (0.1 mol%, 0.5 mol%, and 1 mol%) were evaluated for their cytotoxicity (DOX-loaded liposomes) on MDA-MB-231 (epithelial human breast cancer cells), 4T1 (murine mammary carcinoma cells), and MCF7 (Human breast cancer cells); the two first are TNBC cells and overexpress FA receptor, the third does not. Cytotoxicity results proved that increasing amounts of FA on the surface of liposomes results in enhanced antitumor activity of liposomal-DOX in the case of the cancer cells, which overexpress the FA receptor.

Arsonoliposomes (ARSL) constitute a particular class of liposomes that incorporate arsonolipids (ARS) into their membranes. ARSL realize selective toxicity to cancer cells; thus, they are an important tool in the treatment of cancer. Folic acid (FA) is widely used in targeted drug delivery due to its high affinity for the folate receptors that are overexpressed in cancer cell membranes. The aim of our studies was to develop novel triple-negative breast cancer (TNBC)-targeted ARSL by incorporating folic acid-conjugated polyethylene-glycol PEG-lipid (FA-PEG-lipid) into their membrane and loading them with anticancer drug doxorubicin (DOX). ARSL incorporating 0.1 mol% of FA-PEG-lipid were prepared and loaded with DOX, using the active loading protocol. They were characterized for their size distribution, zeta potential and drug entrapment efficiency (%). Their cytotoxic activity towards TNBC cell lines, particularly MDA-MB-231 (epithelial human breast cancer cells) and MCF7 (human breast cancer cells), was evaluated by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide MTT-assay. The first results demonstrated enhanced toxicity of this novel type of ARSL towards cancer cells, which is particularly interesting and deserves further exploitation.