A Cr2O3-melamine@MAF-6 derived carbon nanozyme composite (CrN@MDC) was developed and integrated into a lateral flow assay (LFA) for the sensitive and specific quantification of transglutaminase 2 (TGM2), a potential biomarker for liver cancer, in clinical plasma samples. CrN(x)@MDC nanozymes with varying Cr2O3 contents (x = 0, 0.05, 0.10, and 0.15 g) were synthesized, and their peroxidase-like activities were evaluated using a 3,3',5,5'-tetramethylbenzidine (TMB)/H2O2 colorimetric reaction, which identified CrN(0.10)@MDC as the most catalytically active composition. The CrN(0.10)@MDC-based LFA generated a distinct black-to-blue colorimetric signal through nanozyme-catalyzed TMB oxidation, enabling detection of TGM2 concentrations as low as 0.025 nM in both buffer and plasma. This corresponded to an approximately 40-fold improvement in visual detection sensitivity compared with a conventional gold nanoparticle-based LFA. In addition to a low limit of detection (0.0095 nM) in a plasma matrix, the assay exhibited high selectivity against abundant plasma proteins, relevant biomarkers, and liver disease-associated interferents, likely attributable to effective nanozyme surface blocking using bovine serum albumin. Direct analysis of undiluted clinical plasma samples from healthy individuals and liver cancer patients showed that TGM2 concentrations measured using the proposed LFA were consistent with those obtained by a commercial enzyme-linked immunosorbent assay. These results support the applicability of the nanozyme-amplified LFA as a proof-of-concept platform for clinically relevant TGM2 detection.

A groundbreaking demonstration of the utilization of the metal-organic framework MIL-101(Fe) as an exceptionally perceptive visual label in colorimetric lateral flow assays (LFA) is described. This pioneering approach enables the precise identification of transglutaminase 2 (TGM2), a recognized biomarker for chronic kidney disease (CKD), in urine specimens, which offers a remarkably sensitive naked-eye detection mechanism. The surface of MIL-101(Fe) was modified with oxalyl chloride, adipoyl chloride, and poly(acrylic) acid (PAA); these not only improved the labeling material stability in a complex matrix but also achieved a systematic control in the detection limit of the TGM2 concentration using our LFA platform. The advanced LFA with the MIL-101(Fe)-PAA label can detect TGM2 concentrations down to 0.012, 0.009, and 0.010 nM in Tris-HCl buffer, urine, and desalted urine, respectively, which are approximately 55-fold lower than those for a conventional AuNP-based LFAs. Aside from rapid TGM2 detection (i.e., within 20 min), the performance of the MIL-101(Fe)-PAA-based LFA on reproducibility [coefficients of variation (CV) < 2.9%] and recovery (95.9-103.2%) along with storage stability within 25 days of observation (CV < 6.0%) shows an acceptable parameter range for quantitative analysis. A sophisticated sensing method grounded in machine learning principles was also developed, specifically aimed at precisely deducing the TGM2 concentration by analyzing immunoreaction sites. More importantly, our developed LFA offers potential for clinical measurement of TGM2 concentration in normal human urine and CKD patients' samples.

A lateral flow assay (LFA) sensor on a half-strip platform labeled with blue carbon nanodot-polystyrene (PS-CND) nanoconjugates was developed for the detection of carcinoembryonic antigen (CEA) levels in buffer and serum solutions from healthy and cancer patients. CNDs, biocompatible nanoparticles containing amino groups synthesized by hydrothermal synthesis, were conjugated to spherical polystyrene (PS) beads with an average diameter of 60 nm, followed by the attachment of a detection probe, anti-CEA (M0911042), using a heterobifunctional cross-linker. PS beads were used as a template in CND conjugates to provide uniform size and shape of fluorescent labels without losing CND fluorescence intensity after the antibody conjugation step and to improve fluorescence stability. Upon the interaction of CEA from samples with the anti-CEA (M0911042) probe-modified PS-CND, which was further adsorbed onto a test line composed of the capture anti-CEA (M0911041) physisorbed onto a nitrocellulose membrane, the fluorescent signals on the test line increased as a function of the CEA concentrations under irradiation with a portable 365 nm ultraviolet lamp. A linear concentration range of 0.04–70 nM in buffer was observed, with a limit of detection of 0.3 nM. The applicability of the developed LFA half-strip sensor for disease diagnosis was demonstrated by identifying fluorescent levels on the test line due to the presence of CEA in serum samples from cancer patients. Importantly, signals from healthy human serum solutions because of lower CEA concentrations beyond the sensor detection capability were clearly distinguished from the patient ones.

A rapid and sensitive method is urgently required for the detection of acute myocardial infarction (AMI), a major cause of death globally. As a user-friendly diagnostic platform, the lateral flow immunoassay (LFIA) has been considered as a rapid and low-cost tool for point-of-care testing. Among the available LFIAs, fluorescent nanoparticles have been widely used as reporters to improve the sensitivity of detection targets. This review describes the basic principle of fluorescence-based LFIA and various fluorescent nanoparticles used as a signal readout. Recently, various studies on fluorescent reporters-based LFIAs for diagnosing potential AMI have been outlined by determining the level of AMI-specific protein biomarkers, including cardiac troponins I and T, heart-type fatty acid-binding protein, myoglobin, brain natriuretic peptide, suppression of tumorigenicity 2, creatine kinase-MB, and C-reactive protein. The challenges of the used fluorescent reporters in LFIA are discussed to achieve affordable and excellent performances for clinical applications.

A lateral flow immunoassay (LFIA) method using carbon nanodot@silica as a signaling material was developed for analyzing the concentration of retinol-binding protein 4 (RBP4), one of the lung cancer biomarkers. Instead of antibodies mainly used as bioreceptors in nitrocellulose membranes in LFIA for protein detection, aptamers that are more economical, easy to store for a long time, and have strong affinities toward specific target proteins were used. A 5' terminal of biotin-modified aptamer specific to RBP4 was first reacted with neutravidin followed by spraying the mixture on the membrane in order to immobilize the aptamer in a porous membrane by the strong binding affinity between biotin and neutravidin. Carbon nanodot@silica nanoparticles with blue fluorescent signal covalently conjugated to the RBP4 antibody, and RBP4 were injected in a lateral flow manner on to the surface bound aptamer to form a sandwich complex. Surfactant concentrations, ionic strength, and additional blocking reagents were added to the running buffer solution to optimize the fluorescent signal off from the sand-wich complex which was correlated to the concentration of RBP4. A 10 mM Tris (pH 7.4) running buffer containing 150 mM NaCl and 0.05% Tween-20 with 0.6 M ethanolamine as a blocking agent showed the optimum assay condition for carbon nanodot@silica-based LFIA. The results indicate that an aptamer, more economical and easier to store for a long time can be used as an alternative immobilizing probe for antibody in a LFIA device which can be used as a point-of-care diagnosis kit for lung cancer diseases.

A fluorescent paper strip immunoassay in conjunction with carbon nanodots@silica (CND@SiO2) as a label was developed for the quantitative measurements of human serum amyloid A1 (hSAA1) in serum at clinically significant concentrations for lung cancer diagnosis. Monodispersed CND@SiO2 was prepared by cohydrolysis between silane-crosslinked carbon nanodots and silica precursors via the Ströber method and further attached covalently to anti-hSAA1 (14F8) monoclonal antibody [anti-hSAA1(14F8)] specific to the hSAA1 target. The hSAA1 concentrations were then determined by quantifying the blue fluorescence intensity upon 365 nm excitation of the captured hSAA1 with anti-hSAA1(14F8)-CND@SiO2 conjugates in the test line on a paper strip where anti-hSAA1 (10G1) monoclonal antibody was physisorbed. The developed fluorescent paper strip with CND@SiO2 can detect hSAA1 at concentrations ranging from 0.1 to 5 nM (R2 = 0.995), with a limit of detection of  0.258 nM in 10 mM phosphate buffer pH 7.4 containing human serum albumin. The performance of  recovery (90.98–109.17%) and repeatability (coefficients of variation < 8.46%) obtained was also acceptable for quantitative determinations. The platform was employed for direct determination of hSAA1 concentrations in undiluted serum samples from lung cancer patients (relative standard deviation (RSD) < 7.46%) and healthy humans (RSD < 3.96%). The results were compared with those obtained using a commercially available enzyme-linked immunosorbent assay alongside liquid chromatography with tandem mass spectrometry measurements.