Alpha-fetoprotein In Serum Samples Research Articles

A photoelectrochemical immunosensing platform for ultrasensitive detection of alpha-fetoprotein based on a signal amplification strategy

For the first time, a PEC immunosensor based on a signal amplification strategy is successfully constructed to quantitatively detect alpha-fetoprotein in serum sample. Three favorable factors explain the ultra-high sensitivity of this method. Firstly, compared with pure BiPO4, the BiPO4/BiOBr heterojunction has a narrower band gap, which expands the light absorption range and enables the light energy to be fully utilized. Secondly, the separation of photogenerated electrons and hole pairs during PEC detection is due to the efficient matching of energy levels among BiPO4, BiOBr and CdS, inhibiting the recombination of photogenerated electrons, which improves the performance of PEC immunosensor. Thirdly, due to the presence of CdS, the light absorption capability of the sensor is enhanced, more electron-hole pairs are generated, and the photocurrent signal is increase. Under the optimal conditions, the PEC immunosensor shows a wide linear range of 0.001–1000 ng/mL for AFP and a low detection limit of 0.82 pg/mL. The PEC immunosensor developed in this experiment has excellent reproducibility, stability and high sensitivity, and also achieves satisfactory results in the analysis of human serum samples, establishing a new analytical method for biomarker detection.

Development of active affibody aggregates induced by a self-assembling peptide for high sensitive detection of alpha-fetoprotein

Alpha-fetoprotein (AFP) is one of the most important biomarkers associated with primary liver cancer and the development of accurate and sensitive methods for detecting the serum level of AFP is of great importance in cancer diagnosis. In this study, three self-assembling peptides (SAPs) tagged AFP specific affibody dimer (ZAFP D2)2 recombinant protein, (ZAFP D2)2-ELK16, (ZAFP D2)2-L6KD and (ZAFP D2)2-R18A were successfully produced and characterized. The (ZAFP D2)2-ELK16 affibody aggregates showed higher and more specific binding affinity to AFP compared to (ZAFP D2)2 and other two SAP-tagged (ZAFP D2)2 and in addition displayed enhanced AFP-detection sensitivity and improved stability. A sensitive two-site enzyme-linked immunosorbent assay for AFP using (ZAFP D2)2-ELK16 as the detection reagent was also established. The AFP detection limit and linear range using (ZAFP D2)2-ELK16 based two-site ELISA were 9.7 ng mL−1 and 20–600 ng mL−1, which were significantly improved compared to those of the (ZAFP D2)2 based two-site ELISA for AFP. To detect AFP in serum samples, the average recovery ranged from 95.02% to 110.41% and the relative standard deviation was in the range of 2.45%-3.97%, which meet the standard clinical testing requirements. This proposed strategy provides a high yield, easy purification and green biosynthesis method to prepare SAP-based affibody aggregates for enhanced detection sensitivity by improving the immunoassay signal amplification activity.

Eu-MOF nanorods functionalized with large heterocyclic ionic liquid for photoelectrochemical immunoassay of α-fetoprotein

An ionic liquid with a sizeable heterocyclic structure was successfully synthesized and used as a linker to coordinate with Eu3+ ions to provide metal-organic framework (Eu-MOF) nanorods. Gold nanoparticles-modified Eu-MOF nanocomposites (Eu-MOF@AuNPs) were synthesized by the in-situ reduction of chloroauric acid on the Eu-MOF nanorod surface. The outstanding conductivity and local surface plasmon resonance (LSPR) of Au-NPs and the distinct nature of MOFs worked together to enhance photoelectrochemical (PEC) performance by decreasing the recombination of photo-excited carriers and taking advantage of a broad absorption range for light harvesting. Under white light irradiation, the Eu-MOF@AuNPs nanocomposite had a better PEC performance than pure Eu-MOF. Therefore, a novel photoelectrochemical immunosensor was fabricated for alpha-fetoprotein (AFP) using Eu-MOF@AuNPs as the photoactive element. When the anti-AFP was attached to the Eu-MOF@AuNPs/GCE interface, the designed immunosensor exhibited a specific photocurrent response to AFP. The specific binding of AFP to anti-AFP can cause a significant steric hindrance, thus hinder the charge transfer and the separation of electron-hole pairs due to large molecular size and poor conductivity. As a result, the PEC immunosensing platform presented a decreasing trend on photocurrent intensity. Under the optimized conditions, the calibration plot showed a linearity with the logarithm of AFP concentration (0.002–15.0 ng mL−1), and a detection limit of 0.16 pg mL−1 was obtained (S/N = 3). Moreover, this immunosensor presented a brilliant selectivity, high reproducibility, and outstanding stability, which possesses remarkable potential applications in determining AFP in serum samples.

An electrodeposited Au nanoparticle/porous graphene nanoribbon composite for electrochemical detection of alpha-fetoprotein

In this work, a novel label-free electrochemical immunosensor for the detection of alpha-fetoprotein (AFP) is fabricated for early diagnosis and prognostics of liver cancer. A porous graphene nanoribbon (PGNR) was synthesized via chemical reduction method and it was electrodeposited with gold nanoparticles (AuNPs) to obtain AuNPs/PGNR hybrid nanomaterial. Anti-AFP was immobilized onto AuNPs/PGNR/glassy carbon electrode and anti-AFP/AuNPs/PGNR/GCE was further studied to demonstrate its electrocatalytic activity towards AFP antigen. PGNR enhances the electroactive surface area and the electron transfer ability between the electrode and redox probe while the AuNPs deposited on PGNR are used to immobilize biomolecules and to facilitate the electron transport. The superior biosensing performance towards AFP under physiological pH condition is demonstrated by a corresponding decreased peak current in differential pulse voltammetry for a wide linear range (5–60 ng/mL) with a low detection limit of 1 ng/mL. Detection of AFP in serum samples by this label-free electrochemical immunosensor without fouling or significant interference implies that the anti-AFP/AuNPs/PGNR modified GCE has a great application potential for clinical diagnosis of AFP.

A novel biosensor for early diagnosis of liver cancer cases using smart nano‐magnetic metal–organic framework

AbstractFor the first time, alpha‐fetoprotein (AFP) quantification as an early test for liver cancer diagnosis competing with other interfering analytes and factors based on a novel nano‐magnetic metal–organic framework (MOF) was investigated. The novel nano‐magnetic MOF was synthesized via a simple preparation method, and characterized using various techniques. The X‐ray photoelectron spectrum of the nano‐magnetic MOF shows Cu 2p, O 1s, N 1s and C 1s peaks that evidence the presence of these elements in the sample without any impurities. Scanning and transmission electron microscopy images show the nano‐magnetic MOF with average size between 22 and 43 nm. The results of a photoluminescence study show that the nano‐magnetic MOF exhibits strong emission at 418 nm after excitation at 354 nm. Moreover, a Stern–Völmer graph shows a linear calibration curve over an AFP concentration range 1.0–520 ng ml−1 with a correlation coefficient of 0.997, detection limit of 1.18 ng ml−1 and quantitation limit of 3.58 ng ml−1. According to the marked quenching of the photoluminescence intensity of the nano‐magnetic MOF using various concentrations of AFP, it was successfully used as a biosensor for AFP in serum samples collected from hepatitis patients in addition to healthy males and females. The quenching mechanism was well studied, and found to be a dynamic type. The present work offers a simple, low‐cost, room‐temperature and rapid technique, being non‐time‐consuming, highly accurate, selective and highly sensitive compared with most published methods.

A novel ECL sensor based on a boronate affinity molecular imprinting technique and functionalized SiO2@CQDs/AuNPs/MPBA nanocomposites for sensitive determination of alpha-fetoprotein

In this work, a boronate-affinity sandwich electrochemiluminescence (ECL) sensor was constructed to detect alpha-fetoprotein (AFP) based on a multiple signal amplification strategy. Gold nanoparticles (AuNPs) were utilized and modified on the surface with chitosan in order to facilitate electron transfer. The composite of the molecularly imprinted polymer (MIP) enhanced the selectivity of alpha-fetoprotein detection. 4-mercaptophenylboronic acid (MPBA) was used as the tracing tag for capture of alpha-fetoprotein. SiO2 nanoparticles carried carbon quantum dots (CQDs) labeled with gold nanoparticles and produced an ECL signal. Under the optimum experimental conditions, the linear range for alpha-fetoprotein was between 0.001 and 1000 ng/mL with a correlation coefficient of 0.9952, and the detection limit was 0.0004 ng/mL (S/N = 3). This proposed ECL sensor displayed several advantages, including outstanding selectivity, fine reproducibility, high sensitivity, low detection limit and wide linear range. Furthermore, the newly constructed boronate-affinity sandwich ECL sensor was successfully applied to the determination of alpha-fetoprotein in serum samples, indicating great potential for application in clinical diagnostics.

Electrochemical Assay of the Alpha Fetoprotein-L3 Isoform Ratio To Improve the Diagnostic Accuracy of Hepatocellular Carcinoma.

Hepatocellular carcinoma (HCC) is now the major malignant disease with high morbidity and mortality, which seriously endangers human lives and health. Alpha fetoprotein (AFP) assay is a commonly used serological biomarker for clinical diagnosis of HCC, but it lacks specificity. Analysis of its isoform AFP-L3, especially the AFP-L3 ratio in total AFP (AFP-L3%), can significantly improve the specificity for HCC identification. Herein, an electrochemical approach has been first proposed for simple, accurate, and fast determination of AFP-L3% in clinical samples. On the basis of two independent electrochemical signals generated from the synthesized nanoparticles, 4-mercaptophenylboronic acid (MPA)-functionalized copper nanoparticles (MPA-CuNPs) and the Lens culinaris agglutinin (LCA)-functionalized silver nanoparticles (LCA-AgNPs), simultaneous quantification of the AFP-L3 and total AFP in serum sample has been achieved, thus achieving directly the electrochemical assay of AFP-L3%. To be noted, both the assay time and the assay procedure have been significantly compressed when compared to that of available techniques in clinical use. Therefore, with the integration of electrochemical techniques, this new approach for AFP-L3% analysis would be promising for the accurate diagnosis of HCC.

Dual-mode fluorescent and colorimetric immunoassay for the ultrasensitive detection of alpha-fetoprotein in serum samples

We present a novel dual-mode fluorescent and colorimetric immunosensor based on conventional immunoassay platforms by utilizing a gold nanoflower (AuNF)-loaded fluorescein molecule (AuNF@Fluorescein) as signal output. The AuNFs were modified with thiolated carboxyl ligand, which consisted of a hydrophobic alkane chain as hydrophobic wallet for fluorescein encapsulation, a tetra (ethylene glycol) unit for biocompatibility and solubility, and a functional carboxyl group for the conjugation of biorecognition molecules for biosensing. The resultant AuNFs showed a high loading capacity of 3.74 × 106 fluorescein molecules per AuNF because of its flower-like shape with many complex branches. By adjusting the solution pH to 8.0, the fluorescein molecules can almost entirely be released from the hydrophobic wallet of AuNF@Fluorescein, which led to strong fluorescent-signal amplification. Under the optimal detection conditions, the proposed immunoassay based on fluorescent signal exhibited ultrahigh sensitivity for alpha-fetoprotein (AFP) detection, with a limit of detection (LOD) of 29 fg/mL. This value is approximately 9.3 × 103-fold lower than that of corresponding horseradish peroxidase (HRP)-based immunoassay (LOD = 270 pg/mL). The fluorescein molecule also had intrinsic peroxidase-like activity to catalyze 3,3′,5,5′-tetramethylbenzidine oxidation with hydrogen peroxide for colorimetric signal. The proposed method with colorimetric mode further exhibited a sensitivity with a LOD of 17.7 pg/mL, which is about 15-fold lower than that of conventional HRP-based immunoassay. The recoveries of the proposed dual-mode immunoassay for AFP spiked serum samples ranged within 89.85%–100.0%, with the coefficient of variations ranging from 0.5% to 2.4%, indicating acceptable accuracy and precision for AFP quantitative detection. The reliability of the developed dual-mode immunoassay was further compared with a commercial chemiluminescence immunoassay kit by analyzing 20 clinical serum samples, showing that the two methods well agreed with each other, with high correlation coefficients of 0.997 and 0.986 based on recorded fluorescence and colorimetric signals, respectively. In summary, the proposed method was highly suitable for the ultrasensitive analysis of biomarkers or infectious diseases by fluorescence mode and can be used for routine clinical diagnosis by colorimetric mode.

Manganese dioxide nanoparticle-based colorimetric immunoassay for the detection of alpha-fetoprotein

An immunoassay has been developed for the determination of alpha-fetoprotein (AFP). It is based (a) on the use of a detection reaction mediated by manganese dioxide nanoparticles (MnO2 NPs) because their good stability at room temperature, and (b) on tyramine signal amplification (TSA). The MnO2 NPs acts as an artificial peroxidase that causes the conversion of TMB to give a colored product, and the tyramine-triggered reaction is used for signal amplification to improve the detection limit. Combined with immuno-magnetic separation and enrichment, the response of this AFP immunoassay is linear from 6.25–400 ng mL−1, with a detection limit of 22 pg mL−1 (S/N = 3). This immunoassay was successfully applied to the quantification of AFP in serum samples, and gave excellent accuracy compared with the clinical results from a local hospital. The LOD and stability of this assay are better than those of the standard horseradish peroxidase-based ELISA. The strategy presented here is conceived to have a wider scope in that it may be extended to various other immunoassays.

A Portable Immunosensor with Differential Pressure Gauges Readout for Alpha Fetoprotein Detection

A portable, affordable and simple detector is requested in a “Point-of-Care-Testing” (POCT) system. In this study, we exploited the potentialities of Differential Pressure Gauge (DPG) to the orientation of POCT technology. Alpha fetoprotein (AFP) was chosen as a model analyte that could specifically recognized by its antigen, and a tiny outfits equipped with a DPG was employed as the signal readout. Pt/SiO2 nanospheres were synthesized and modified with the detection antibody. In the presence of target, a sandwich of immunocomplex specifically formed and the Pt/SiO2 had been modified on the capture antibody. Which then can be dissolved to release plenty of Pt and the suspensions were transferred into a closed vial filled with appropriated amount of hydrogen peroxide. Subsequently, hydrogen peroxide was decomposed to produce oxygen, resulting in the enhancement of pressure in the closed vial and which can be detected by DPG easily. Under the optimized conditions, the read out signal from DPG had a direct relationship with AFP concentrations in the range of 10~200 ng/mL, and the detection limit was as low as 3.4 ng/mL. The proposed portable sensor had been successfully applied to detect AFP in serum samples with satisfactory results. This strategy holds a great promising in biological analysis as its convenient operations, reliable results and flexible apparatus.

Prussian blue–gold nanoparticles-ionic liquid functionalized reduced graphene oxide nanocomposite as label for ultrasensitive electrochemical immunoassay of alpha-fetoprotein

In this work, poly(diallyldimethylammonium chloride) (PDDA) protected Prussian blue/gold nanoparticles/ionic liquid functionalized reduced graphene oxide (IL-rGO-Au-PDDA-PB) nanocomposite was fabricated. The resulting nanocomposite exhibited high biocompatibility, conductivity and catalytic activity. To assess the performance of the nanocomposite, a sensitive sandwich-type immunosensor was constructed for detecting alpha-fetoprotein (AFP). Greatly enhanced sensitivity for this immunosensor was based on triple signal amplification strategies. Firstly, IL-rGO modified electrode was used as biosensor platform to capture a large amount of antibody due to its increased surface area, thus amplifying the detection response. Secondly, a large number of Au-PDDA-PB was conjugated on the surface of IL-rGO, which meant the enrichment of the signal and the more immobilization of label antibody. Finally, the catalytic reaction between H2O2 and the IL-rGO-Au-PDDA-PB nanocomposite further enhanced the signal response. The signals increased linearly with AFP concentrations in the range of 0.01–100ngmL−1. The detection limit for AFP was 4.6pgmL−1. The immunosensor showed high sensitivity, excellent selectivity and good stability. Moreover, the immunosensor was applied to the analysis of AFP in serum sample with satisfactory result.

Sensitive sandwich electrochemical immunosensor for alpha fetoprotein based on prussian blue modified hydroxyapatite

A sandwich electrochemical immunosensor for the sensitive determination of alpha fetoprotein (AFP) has been fabricated. Prussian blue modified hydroxyapatite (PB@HAP) was firstly prepared and used as electrochemical label due to the wonderful conductivity and good biocompatibility of HAP. The results proved that the immunosensor fabricated using the label based on PB@HAP loaded with horse radish peroxidase (HRP) and secondary anti-AFP antibody (Ab2) (PB@HAP-HRP-Ab2) had high sensitivity, and the sensitivity of the label PB@HAP-HRP-Ab2 was much higher than labels of PB@HAP-Ab2, PB-HRP-Ab2 and HAP-HRP-Ab2. The mixture of graphene sheet (GS) and thionine (TH) was not only used to immobilize anti-AFP antibody (Ab1) but also took part in the signal amplification. The amperometric signal increased linearly with AFP concentration in the range of 0.02–8ng/mL with a low detection limit of 9pg/mL. The immunosensor had the advantages of high sensitivity, good selectivity and good stability, and was applied to the analysis of AFP in serum sample with satisfactory results. Due to the low-cost and easy synthesis of PB@HAP, the screen-printed electrodes could be used instead of the bare glass carbon electrode in order to achieve mass production. In addition, it had potential application in the detection of other tumor markers.

A novel label-free electrochemical immunosensor based on graphene and thionine nanocomposite

A novel label-free electrochemical immunosensor for the detection of alpha fetoprotein (AFP) has been developed by immobilizing anti-AFP on graphene sheet (GS) and thionine (TH) modified glassy carbon electrode. The modification procedure was characterized by cyclic voltammetry (CV). The detection is based on the peak current change of the modified electrode before and after the antigen–antibody reaction. Tests relating to the detection of AFP demonstrated that the immunosensor exhibited high sensitivity, wide linear range (0.05–2.00 ng mL −1), low detection limit (5.77 pg mL −1 based S/ N = 3) and good long-term stability. The reliability of the developed immunosensor has been proved by the detection of AFP in serum samples with satisfactory results.

Differential reactivity of alpha-fetoprotein with lectins and evaluation of its usefulness in the diagnosis of hepatocellular carcinoma.

The reactivity of alpha-fetoprotein (AFP) with concanavalin A (Con A) and Lens culinaris agglutinin (LCA) was studied by crossed immuno-affinoelectrophoresis of the serum samples of 146 patients from three groups (groups I, II and III). Fifty-one patients with benign liver diseases were included in group I, 83 patients with hepatocellular carcinoma in group II, and 12 patients with carcinoma metastatic to the liver from digestive organs in group III. In group I, the percentage of Con A-reactive species of AFP was high (97 +/- 5%, mean +/- SD), but that of LCA-reactive species was very low (3 +/- 5%). The percentage of LCA-reactive species of AFP in group II (45 +/- 33%) was higher than that in group I (P less than 0.001), while the Con A binding pattern of AFP in this group, as demonstrated by immuno-affinoelectrophoresis, was similar to that of group I. The percentage of Con A-reactive species of AFP in group III (55 +/- 24%) was much lower than that in group II (97 +/- 5%) (P less than 0.001). The above results indicate that measurement of the reactivity of AFP in serum samples with Con A and LCA is useful for the differentiation of AFP species which are found in association with benign liver diseases, hepatocellular carcinoma and carcinoma metastatic to the liver.

Differences in the structure of alpha-fetoprotein and its clinical use in pediatric surgery

Techniques have been studied which distinguish two variants of human alpha-fetoprotein (AFP) on the basis of characteristics of the carbohydrate moiety of this glycoprotein. AFP in serum samples from six children with tumors of yolk sac origin showed little concanavalin-A (Con A) binding. In contrast, Con A binding of AFP was almost complete in serum samples from 14 other subjects with elevated AFP, including two with liver-cell tumors, eight with neonatal cholestasis, and four normal newborn infants. Differences were confirmed by immunoelectrophoretic studies. Thus, AFP from cells of yolk sac origin can be distinguished from AFP from liver cells or from tumors of hepatic cell origin.

Alpha-fetoprotein (AFP) levels in maternal serum and amniotic fluid in singleton pregnant women in their 10th-25th week post last menstrual period.

A rapid paper disk radioimmunoassay of the sandwich type was used to determine the levels of alpha fetoprotein (AFP) in 826 maternal serum samples derived from an equal number of singleton pregnant women prospectively investigated during their 10th-25th week past last menstrual period (LMP). In addition 396 amniotic fluid (AF) samples from the 14th-20th week post LMP were analyzed. Theoretical ranges of normal AFP variation are presented. The extreme range of AFP in serum samples from 236 nonpregnant blood donors varied fron .2-18.3 mcg/liter. Variability of AFP in maternal serum samples increased considerably with gestational progress. A specific low cut-off could not be defined until Week 15 post LMP because of the overlap between distribution of AFP levels in the pregnant and nonpregnant populations studied. However from Week 15 post LMP a feasibly low and decreasing frequency of maternal serum samples from normal pregnancies still had an AFP value below 10 mcg/liter; this value also represented the upper limit for 98.5% of the nonpregnant population. Thus 10 mcg/liter was found to be a useful cut-off from the 15th post LMP week below which pregnancy complications or severe gestation age miscalculations could be deduced. For AF AFP values a strong negative correlation between gestational age and AFP levels was found at least up to Week 20 post LMP.