Search Results (20)

Mycotoxins are responsible for a multitude of diseases in both humans and animals, resulting in significant medical and economic burdens worldwide. Conventional detection methods, such as enzyme-linked immunosorbent assay (ELISA), high-performance liquid chromatography (HPLC), and liquid chromatography-tandem mass spectrometry (LC-MS/MS), are highly effective, but they are generally confined to laboratory settings. Consequently, there is a growing demand for point-of-care testing (POCT) solutions that are rapid, sensitive, portable, and cost-effective. Lateral flow assays (LFAs) are a pivotal technology in POCT due to their simplicity, rapidity, and ease of use. This review synthesizes data from 78 peer-reviewed studies published between 2015 and 2024, evaluating advances in nanoparticle-based LFAs for detection of singular or multiplex mycotoxin types. Gold nanoparticles (AuNPs) remain the most widely used, due to their favorable optical and surface chemistry; however, significant progress has also been made with silver nanoparticles (AgNPs), magnetic nanoparticles, quantum dots (QDs), nanozymes, and hybrid nanostructures. The integration of multifunctional nanomaterials has enhanced assay sensitivity, specificity, and operational usability, with innovations including smartphone-based readers, signal amplification strategies, and supplementary technologies such as surface-enhanced Raman spectroscopy (SERS). While most singular LFAs achieved moderate sensitivity (0.001–1 ng/mL), only 6% reached ultra-sensitive detection (<0.001 ng/mL), and no significant improvement was evident over time (ρ = −0.162, p = 0.261). In contrast, multiplex assays demonstrated clear performance gains post-2022 (ρ = −0.357, p = 0.0008), largely driven by system-level optimization and advanced nanomaterials. Importantly, the type of sample matrix (e.g., cereals, dairy, feed) did not significantly influence the analytical sensitivity of singular or multiplex lateral LFAs (Kruskal–Wallis p > 0.05), confirming the matrix-independence of these optimized platforms. While analytical challenges remain for complex targets like fumonisins and deoxynivalenol (DON), ongoing innovations in signal amplification, biorecognition chemistry, and assay standardization are driving LFAs toward becoming reliable, ultra-sensitive, and field-deployable platforms for high-throughput mycotoxin screening in global food safety surveillance. Full article

Background: Acute Kidney Injury (AKI) is a condition that affects a significant proportion of acutely unwell patients and is associated with a high mortality rate. Patients undergoing haemopoietic stem cell transplantation (HSCT) are in an extremely high group for AKI. Identifying a biomarker or panel of markers that can reliably identify at-risk individuals undergoing HSCT can potentially impact management and outcomes. Early identification of AKI can reduce its severity and improve prognosis. We evaluated the urinary Liver type fatty acid binding protein (L-FABP), a tubular stress and injury biomarker both as an ELISA and a point of care (POC) assay for AKI detection in HSCT. Methods: 85 patients that had undergone autologous and allogenic HSCT (35 and 50, respectively) had urinary L-FABP (uL-FABP) measured by means of a quantitative ELISA and a semi-quantitative POC at baseline, day 0 and 7 post-transplantation. Serum creatinine (SCr) was also measured at the same time. Patients were followed up for 30 days for the occurrence of AKI and up to 18 months for mortality. The sensitivity and specificity of uL-FABP as an AKI biomarker were evaluated and compared to the performance of sCr using ROC curve analysis and logistic regression. Results: 39% of participants developed AKI within 1 month of their transplantation. The incidence of AKI was higher in the allogenic group than in the autologous HTSC group (57% vs. 26%, p = 0.008) with the median time to AKI being 25 [range 9-30] days. This group was younger (median age 59 vs. 63, p < 0.001) with a lower percentage of multiple myeloma as the primary diagnosis (6% vs. 88%, p < 0.001). The median time to AKI diagnosis was 25 [range 9–30] days. uL-FABP (mcg/gCr) at baseline, day of transplant and on the 7th day post-transplant were 1.61, 5.39 and 10.27, respectively, for the allogenic group and 0.58, 4.36 and 5.14 for the autologous group. Both SCr and uL-FABP levels rose from baseline to day 7 post-transplantation, while the AUC for predicting AKI for baseline, day 0 and day 7 post-transplant was 0.54, 0.59 and 0.62 for SCr and for 0.49, 0.43 and 0.49 uL-FABP, respectively. Univariate logistic regression showed the risk of AKI to be increased in patients with allogenic HSCT (p = 0.004, 95%CI [0.1; 0.65]) and in those with impaired renal function at baseline (p = 0.01, 95%CI [0.02, 0.54]). The risk of AKI was also significantly associated with SCr levels on day 7 post-transplant (p = 0.03, 95%CI [1; 1.03]). Multivariate logistic regression showed the type of HSCT to be the strongest predictor of AKI at all time points, while SCr levels at days 0 and 7 also correlated with increased risk in the model that included uL-FABP levels at the corresponding time points. The POC device for uL-FABP measurement correlated with ELISA (p < 0.001, Spearman ‘correlation’ = 0.54) Conclusions: The urinary biomarker uL-FABP did not demonstrate an independent predictive value in the detection of AKI at all stages. The most powerful risk predictor of AKI in this setting appears to be allograft recipients and baseline renal impairment, highlighting the importance of clinical risk stratification. Urinary L-FAPB as a POC biomarker was comparable to ELISA, which provides an opportunity for simple and rapid testing. However, the utility of LFABP in AKI is unclear and needs further exploration. Whether screening through rapid testing of uL-FABP can prevent or reduce AKI severity is unknown and merits further studies. Full article

Herein, we present a novel approach to quantify ferritin based on the integration of an Enzyme-Linked Immunosorbent Assay (ELISA) protocol on a Graphene Field-Effect Transistor (gFET) for bioelectronic immunosensing. The G-ELISA strategy takes advantage of the gFET inherent capability of detecting pH changes for the amplification of ferritin detection using urease as a reporter enzyme, which catalyzes the hydrolysis of urea generating a local pH increment. A portable field-effect transistor reader and electrolyte-gated gFET arrangement are employed, enabling their operation in aqueous conditions at low potentials, which is crucial for effective biological sample detection. The graphene surface is functionalized with monoclonal anti-ferritin antibodies, along with an antifouling agent, to enhance the assay specificity and sensitivity. Markedly, G-ELISA exhibits outstanding sensing performance, reaching a lower limit of detection (LOD) and higher sensitivity in ferritin quantification than unamplified gFETs. Additionally, they offer rapid detection, capable of measuring ferritin concentrations in approximately 50 min. Because of the capacity of transistor miniaturization, our innovative G-ELISA approach holds promise for the portable bioelectronic detection of multiple biomarkers using a small amount of the sample, which would be a great advancement in point–of–care testing. Full article

The first point-of-care (PoC) test (v-RetroFel^®; modified version 2021) determining the presence of FeLV p27 antigen and FeLV anti-p15E antibodies has become recently commercially available to identify different feline leukaemia virus (FeLV) infection outcomes. This study aimed to assess this PoC test’s performance concerning FeLV p27 antigen and FeLV anti-p15E antibody detection. Sensitivity, specificity, positive and negative predictive values (PPV, NPV) were assessed after ten minutes (recommended) and 20 min (prolonged) incubation times. The test results were evaluated as either positive or negative. Serum samples from 934 cats were included, originating from Italy (n = 269), Portugal (n = 240), Germany (n = 318), and France (n = 107). FeLV p27 antigen and anti-p15E antibodies were measured by reference standard ELISAs and compared to the PoC test results. The PoC test was easy to perform and the results easy to interpret. Sensitivity and specificity for FeLV p27 antigen were 82.8% (PPV: 57.8%) and 96.0% (NPV: 98.8%) after both, ten and 20 minues of incubation time. Sensitivity and specificity for anti-p15E antibodies were 31.4% (PPV: 71.6%) and 96.9% (NPV: 85.1%) after ten minutes incubation time; sensitivity was improved by a prolonged incubation time (20 min) to 40.0% (PPV: 76.3%), while specificity remained the same (96.9%, NPV: 86.7%). Despite the improved sensitivity using the prolonged incubation time, lower than ideal sensitivities for both p27 antigen and especially anti-p15E antibodies were found, indicating that the PoC test in its current version needs further improvement prior to application in the field. Full article

The aim of this study was to investigate the coinfection of feline retroviruses (feline immunodeficiency virus—FIV, and the feline leukemia virus—FeLV) with Leishmania infantum and Toxoplasma gondii and the factors associated with these pathogens in domestic cats from Mossoró, a city endemic for canine and human leishmaniasis situated in the semiarid region of Northeast Brazil. Blood samples from 120 cats were collected, and an epidemiological questionnaire was applied to investigate the risk factors associated with the infections. Retroviruses, L. infantum, and T. gondii infections were assessed using a point-of-care ELISA and quantitative PCR (qPCR), indirect fluorescent antibody test (IFAT) and qPCR, and IFAT, respectively. The overall seroprevalences observed were 35% (95% CI = 27.0–43.8%) for FIV, 0.8% (95% CI = 0.1–4.5%) for FeLV, 25.8% (95% CI = 18.8–34.3%) for T. gondii, and 4.2% (95% CI = 1.7–9.3%) for L. infantum. Coinfection with FIV and L. infantum was observed in 2.5% (3/120) of the assessed cats, while 12.5% (15/120) were coinfected with FIV and T. gondii. No significant association was found among the investigated agents (p > 0.05). The factors associated with FIV infection in the multivariable analysis were male sex and age above 78 months. The findings of this study demonstrated a high rate of FIV infection in cats from the Brazilian semiarid region and the exposure of these animals to zoonotic and opportunistic agents. Due to the immunosuppressive potential of FIV, cats infected with this retrovirus should be screened for coinfections with L. infantum and T. gondii, and preventative measures should be adopted. Full article

The introduction of point-of-care (POC) assays into clinical practice in patients with inflammatory disease enables on-demand therapeutic decision making. The aim of this study was to compare the POC test Quantum blue (Bühlmann Laboratories) for infliximab (IFX), adalimumab (ADL), and its anti-drug antibodies with the traditional ELISA assay (Promonitor). A total of 200 serum samples were analyzed. Samples were classified into the following three different groups; sub-therapeutic range (IFX < 3 μg/mL and ADL < 5 μg/mL); therapeutic range (IFX: 3–7 μg/mL and ADL: 5–12 μg/mL) and supra-therapeutic range (IFX levels > 7 μg/mL and ADL levels > 12 μg/mL). Significant higher values were measured using the POC test (p < 0.001) for IFX results but no differences in ADL trough levels were observed (p = 0.3101). Spearman’s correlation indicated a good correlation between the two assays (rs = 0.88 for ADL and rs = 0.93 for IFX), and McNemar’s test revealed significant differences (p = 0.016) when classifying IFX samples between therapeutic and supra-therapeutic ranges but no significant differences were found among the other ranges for either IFX or ADL. These results show that we should be cautious when using these rapid measurement methods, and new targets should probably be defined for IFX when using this new analytical method. Full article

Highly active antiretroviral therapy (HAART) includes very potent drugs that are often characterized by high toxicity. Tenofovir (TFV) is a widely used drug prescribed mainly for pre-exposure prophylaxis (PreP) and the treatment of human immunodeficiency virus (HIV). The therapeutic range of TFV is narrow, and adverse effects occur with both underdose and overdose. The main factor contributing to therapeutic failure is the improper management of TFV, which may be caused by low compliance or patient variability. An important tool to prevent inappropriate administration is therapeutic drug monitoring (TDM) of compliance-relevant concentrations (ARCs) of TFV. TDM is performed routinely using time-consuming and expensive chromatographic methods coupled with mass spectrometry. Immunoassays, such as enzyme-linked immunosorbent assays (ELISAs) and lateral flow immunoassays (LFIAs), are based on antibody–antigen specific recognition and represent key tools for real-time quantitative and qualitative screening for point-of-care testing (POCT). Since saliva is a non-invasive and non-infectious biological sample, it is well-suited for TDM. However, saliva is expected to have a very low ARC for TFV, so tests with high sensitivity are required. Here, we have developed and validated a highly sensitive ELISA (IC50 1.2 ng/mL, dynamic range 0.4–10 ng/mL) that allows the quantification of TFV in saliva at ARCs and an extremely sensitive LFIA (visual LOD 0.5 ng/mL) that is able to distinguish between optimal and suboptimal ARCs of TFV in untreated saliva. Full article

Different feline leukemia virus (FeLV) infection outcomes are possible in cats following natural exposure, such as progressive infections (persistent viremia), regressive infections (transient or no viremia followed by proviral persistence) and abortive infections (presence of only antibodies). Laboratory-based testing is currently required for categorization of infection outcomes in cats. The aim of this study was to evaluate the field performance of a novel, rapid, combination point-of-care (PoC) test kit commercially available in Europe (v-RetroFel^®Ag/Ab; 2020–2021 version) to determine different FeLV infection outcomes by concurrent detection of FeLV antigen (p27) and antibodies against FeLV transmembrane envelope protein (p15E). A secondary aim was to evaluate the performance of the same test kit (v-RetroFel^®FIV) to determine positive/negative feline immunodeficiency virus (FIV) infection status by the detection of antibodies to FIV capsid protein (p24) and transmembrane glycoprotein (gp40). Two cohorts of domestic cats were recruited and tested with v-RetroFel^® using plasma or serum, including cats in Australia (n = 200) and cats in Germany (n = 170). Results from p27 antigen PoC testing, proviral DNA PCR, and neutralizing antibody testing or testing for antibodies against non-glycosylated surface unit envelope protein (p45) were used to assign cats to groups according to different FeLV infection outcomes. Testing with a laboratory-based FeLV p15E antibody ELISA was also performed for comparison. In the first cohort, v-RetroFel^®Ag/Ab correctly identified 89% (109/122) FeLV-unexposed cats and 91% (21/23) progressive infections, but no regressive (0/23) or abortive (0/32) infections. In the second cohort, v-RetroFel^®Ag/Ab correctly identified 94% (148/158) FeLV-unexposed cats and 100% (4/4) progressive infections, but no regressive (0/2) and only 17% (1/6) abortive infections. There was test agreement between v-RetroFel^®Ab and the p15E laboratory ELISA in 58.9% of samples. As a secondary outcome of this study, the sensitivity and specificity of v-RetroFel^®FIV testing in cohort 1 were 94.7% (18/19) and 98.3% (178/181), and in cohort 2, 30.0% (3/10) and 100.0% (160/160), respectively. Prior history of FIV vaccination did not produce any false-positive FIV results. In conclusion, v-RetroFel^®Ag/Ab (2020–2021 version) was unable to accurately determine different FeLV infection outcomes in the field. Improvements of the test prior to application to field samples are required. Full article

The detection of vaccine-induced HIV antibody responses by rapid diagnostic tests (RDTs) may confound the interpretation of HIV testing results. We assessed the impact of vaccine-induced seroreactivity (VISR) on the diagnosis of HIV in sub-Saharan Africa. Samples collected from healthy participants of HIVIS and TaMoVac HIV vaccine trials after the final vaccination were analyzed for VISR using HIV testing algorithms used in Mozambique and Tanzania that employ two sequential RDTs. The samples were also tested for VISR using Enzygnost HIV Integral 4 ELISA and HIV western blot assays. Antibody titers to subtype C gp140 were determined using an in-house enzyme-linked immunosorbent assay (ELISA). The frequency of VISR was 93.4% (128/137) by Enzygnost HIV Integral 4 ELISA, and 66.4% (91/137) by western blot assay (WHO interpretation). The proportion of vaccine recipients that would have been misdiagnosed as HIV-positive in Mozambique was half of that in Tanzania: 26.3% (36/137) and 54.0% (74/137), respectively, p < 0.0001. In conclusion, the HIV RDTs and algorithms assessed here will potentially misclassify a large proportion of the HIV vaccine recipients if no other test is used. Increased efforts are needed to develop differential serological or molecular tools for use at the point of care. Full article

Endometriosis or adenomyosis can be clinically diagnosed by ultrasound, symptoms, physical examination, and serum CA125. The urinary markers need to be investigated. The aim of our study was to investigate the urinary markers of clinical endometriosis/adenomyosis, and the correlation of serum CA125 was also studied. From the literature, alpha-1 antitrypsin (A1AT), enolase-1, vitamin D binding protein (VDBP), and CA125 in urine and serum were used in our study and measured by enzyme-linked immunosorbent assays (ELISA). Further clinical correlation and detection performance were evaluated. We enrolled 19 normal controls and 33 patients clinically diagnosed with endometriosis/adenomyosis. There were significant differences between studied patients and normal controls, as follows: serum CA125 (130.91 vs. 19.75 U/mL, p = 0.004); urinary CA125-creatinine ratio (5.591 vs. 0.254 ng/mg, p = 0.028); and urinary VDBP-creatinine ratio (28.028 vs. 7.301 ng/mg, p = 0.018). For diagnostic performances, serum CA125 provided the best results, with an area under curve (AUC) of 0.888 (p = 0.001) and accuracy of 86.5%. Other excellent results were also found using urinary VDBP (AUC 0.841, p = 0.001) and A1AT (AUC 0.722, p = 0.011) creatinine ratio. Using three combined biomarkers, serum CA125, urinary VDBP, and A1AT creatinine ratio, provided good detection power (AUC 0.913, p = 0.001, sensitivity 90.9%, specificity 76.5%). Double urine markers used in combination with VDBP and A1AT creatinine ratio also provided good diagnostic performance (AUC 0.809, p = 0.001, sensitivity 81.8%, specificity 76.5%, accuracy 80%). Further development of non-invasive point-of-care tests using these biomarkers could be a fruitful future endeavor. Full article

To evaluate point-of-care testing (POCT) for the potential early detection of biomarkers of Parkinson’s disease, a systematic investigation of portable and low-cost platforms is performed based on the Proton-enzyme-linked immunosorbent assay (Proton-ELISA) methodology. The detection of the α-synuclein antigen was first presented by biotin-relative linkers, and glucose substrate solution was first performed with a systematic experimental design to optimize the sensing results. All materials in this study are commercially available. Three different experiments with the partitional check were performed to investigate the Proton-ELISA platform, including proton catalyzed efficiency, blocking efficiency, and full Proton-ELISA procedure. The response time was selected as 15 min by the time-dependent curves of a full reaction. The limit of detection of conventional ELISA kits is 0.169 ng/mL, which is much lower than the Proton-ELISA results. The final response of the full Proton-ELISA procedure to pH changes was approximately 0.60 and 0.12 for α-synuclein antigen concentrations of 100 ng/mL and 4 ng/mL, respectively. With the partitional check, pH changes of pure glucose substrate and conjugated oxidase and interference of the nonspecific binding are 1.7 and 0.04, respectively. The lower pH changes far from the partitional check results can be concluded for the properties of glucose oxidase conjugation, including the isoelectric point and binding affinity modification by the testing environment. This preliminary guideline can be used as a lesson learnt to speed up following studies of the evaluation and optimization of other antigen detection. Therefore, Proton-ELISA can be suggested for some special applications with the help of custom-designed conjugation in the environment with less degradation or interference and a proper detection concentration range. Full article

Efforts to control SARS-CoV-2 have been challenged by the emergence of variant strains that have important implications for clinical and epidemiological decision making. Four variants of concern (VOCs) have been designated by the Centers for Disease Control and Prevention (CDC), namely, B.1.617.2 (delta), B.1.1.7 (alpha), B.1.351 (beta), and P.1 (gamma), although the last three have been downgraded to variants being monitored (VBMs). VOCs and VBMs have shown increased transmissibility and/or disease severity, resistance to convalescent SARS-CoV-2 immunity and antibody therapeutics, and the potential to evade diagnostic detection. Methods are needed for point-of-care (POC) testing to rapidly identify these variants, protect vulnerable populations, and improve surveillance. Antigen-detection rapid diagnostic tests (Ag-RDTs) are ideal for POC use, but Ag-RDTs that recognize specific variants have not yet been implemented. Here, we describe a mAb (2E8) that is specific for the SARS-CoV-2 spike protein N501 residue. The 2E8 mAb can distinguish the delta VOC from variants with the N501Y meta-signature, which is characterized by convergent mutations that contribute to increased virulence and evasion of host immunity. Among the N501Y-containing mutants formerly designated as VOCs (alpha, beta, and gamma), a previously described mAb, CB6, can distinguish beta from alpha and gamma. When used in a sandwich ELISA, these mAbs sort these important SARS-CoV-2 variants into three diagnostic categories, namely, (1) delta, (2) alpha or gamma, and (3) beta. As delta is currently the predominant variant globally, they will be useful for POC testing to identify N501Y meta-signature variants, protect individuals in high-risk settings, and help detect epidemiological shifts among SARS-CoV-2 variants. Full article

Periodontitis is a host-mediated bacterial disease that affects the tooth attachment apparatus. Metalloproteinase-8 (MMP-8), a validated biomarker, could aid in clinical diagnosis. This study aimed to evaluate the diagnostic performance of active (a) MMP-8 immunotest versus total (t) MMP-8 ELISA for quantitative real-time diagnosis and assessment of periodontitis severity at the site level. Gingival crevicular fluid (GCF) was sampled from 30 healthy, 42 mild, and 59 severe periodontitis sites from thirty-one volunteers. MMP-8 concentrations were determined by time-resolved immunofluorometric assay (IFMA) and enzyme-linked immunosorbent assay (ELISA). Statistical analysis was performed using the STATA package. Both active and total MMP-8-based methods discriminated among sites according to periodontal diagnosis and severity, with a positive correlation between the two tests (p < 0.001). (a) MMP-8 models showed the best performance in receiver operating characteristic (ROC) curves to discriminate between healthy and periodontitis sites (area under the curve [AUC] = 0.89), while (t) MMP-8 demonstrated a high diagnostic precision in the detection of mild from severe periodontitis sites (AUC ≥ 0.80). The use of (a) MMP-8 and (t) MMP-8 could represent a useful adjunctive tool for periodontitis diagnosis and severity. These results support the applicability of new point-of-care methods in the monitoring of high-risk periodontal patients. Full article

Although the antibody response induced by primary vaccination with Fel-O-Vax^® FIV (three doses, 2–4 weeks apart) is well described, the antibody response induced by annual vaccination with Fel-O-Vax^® FIV (single dose every 12 months after primary vaccination) and how it compares to the primary antibody response has not been studied. Residual blood samples from a primary FIV vaccination study (n = 11), and blood samples from cats given an annual FIV vaccination (n = 10), were utilized. Samples from all 21 cats were tested with a commercially available PCR assay (FIV RealPCR^TM), an anti-p24 microsphere immunoassay (MIA), an anti-FIV transmembrane (TM; gp40) peptide ELISA, and a range of commercially available point-of-care (PoC) FIV antibody kits. PCR testing confirmed all 21 cats to be FIV-uninfected for the duration of this study. Results from MIA and ELISA testing showed that both vaccination regimes induced significant antibody responses against p24 and gp40, and both anti-p24 and anti-gp40 antibodies were variably present 12 months after FIV vaccination. The magnitude of the antibody response against both p24 and gp40 was significantly higher in the primary FIV vaccination group than in the annual FIV vaccination group. The differences in prime versus recall post-vaccinal antibody levels correlated with FIV PoC kit performance. Two FIV PoC kits that detect antibodies against gp40, namely Witness^® and Anigen Rapid^®, showed 100% specificity in cats recently administered an annual FIV vaccination, demonstrating that they can be used to accurately distinguish vaccination and infection in annually vaccinated cats. A third FIV PoC kit, SNAP^® Combo, had 0% specificity in annually FIV-vaccinated cats, and should not be used in any cat with a possible history of FIV vaccination. This study outlines the antibody response to inactivated Fel-O-Vax^® FIV whole-virus vaccine, and demonstrates how best to diagnose FIV infection in jurisdictions where FIV vaccination is practiced. Full article

Longitudinal studies of cats naturally infected with feline leukemia virus (FeLV) are important for understanding disease outcomes. Levels of p27 antigen and copy numbers of proviral DNA have been associated with FeLV-infection courses. The purpose of this prospective study was to establish cutoff values for p27 antigen concentration and proviral DNA load that distinguished high positive from low positive groups of cats and to evaluate an association with survival. At enrollment, 254 cats were tested by point-of-care and microtiter plate enzyme-linked immunosorbent assays (ELISAs) for p27 antigen and real-time polymerase chain reaction (PCR) for proviral DNA. The 127 positive cats were retested monthly for six months and monitored for survival over the four-year study. A receiver operating characteristic-based analysis of samples with concordant or discordant qualitative results for p27 antigen and proviral DNA was used to establish cutoff values, and when applied to test results at enrollment for classifying cats as high positive or low positive, a significant difference in survival was observed. High positive cats had a median survival of 1.37 years (95% CI 0.83–2.02) from time of enrollment, while most low positive cats were still alive (93.1% survival). Quantitative results for p27 antigen concentration and proviral DNA load were highly correlated with survival times in FeLV-infected cats. Full article