Detection Of Human Epididymis Protein Research Articles

A signal-on electrochemical aptasensor for sensitive detection of human epididymis protein 4 based on functionalized metal–organic framework/ketjen black nanocomposite

A signal-on electrochemical aptasensor for sensitive detection of human epididymis protein 4 based on functionalized metal–organic framework/ketjen black nanocomposite

Label-free electrochemical bioplatform based on Au-modified magnetic Fe3O4/α-Fe2O3 hetero-nanorods for sensitive quantification of ovarian cancer tumor marker

Due to challenges in the ultrasensitive detection of human epididymis protein 4 (HE4), a vital tumor marker for the early diagnosis of ovarian cancer (OC), it is necessary to establish efficient methods for HE4 determination. Recently, electrochemical bioplatforms combined with nanomaterials have shown outstanding performance in the analysis of biological molecules. The surface modification of conductive materials is essential for signal amplification and detection sensitivity. Herein, we developed a novel label-free HE4 electrochemical bioplatform by introducing magnetic Fe3O4/α-Fe2O3 hetero-nanorods (Fe3O4/α-Fe2O3 NRs) as a substrate for signal amplification and DNA aptamer (Apt) probe immobilization in combination with magnetic self-assembly and magnetic separation. Differential pulse voltammetry (DPV) results verified that the platform had excellent specificity for HE4 under optimized conditions and was positively linearly correlated with the logarithm of HE4 concentration (R2 = 0.996) with a linear range of 0.1 pg mL−1 to 1 ng mL−1 and detection limit (LOD) of 27.5 fg mL−1. Moreover, the bioplatform demonstrated promising results for the detection of HE4 in spiked serum samples.

Black phosphorus nanosheets/poly(allylamine hydrochloride) based electrochemical immunosensor for the selective detection of human epididymis protein 4

In this work, an electrochemical immunosensor based on black phosphorus nanosheets (BPNS)/poly(allylamine hydrochloride) (PAH) nanocomposite modified glassy carbon electrode was developed for the detection of ovarian cancer biomarker HE4. PAH has been applied to retain BPNS in its original honeycomb structure and to anchor biomolecules electrostatically on the transducer surface. The as synthesized nanocomposite was characterized by zeta potential analysis, scanning electron microscopy, x-ray photoelectron spectroscopy, transmission electron microscopy, high-resolution transmission electron microscopy. Subsequently, the performance of the electrochemical immunosensor was evaluated through cyclic voltammetry, differential pulse voltammetry and electrochemical impedance spectroscopy. Under the optimal condition, the developed electrochemical immunosensor permitted to detect HE4 with a linear range of 0.1–300 ng ml−1 and a detection limit of 0.01 ng ml−1. The developed sensor exhibited good selectivity and specificity to HE4 with negligible interference effect from common biomolecules like bovine serum albumin, lysozyme, protamine, glucose, fructose, hemoglobin and fetal bovine serum. Further, practical application of developed electrochemical immunosensor was demonstrated in spiked human serum which showed satisfactory recovery percentages.

Fluorescence turn-on immunosensing of HE4 biomarker and ovarian cancer cells based on target-triggered metal-enhanced fluorescence of carbon dots

Rapid and sensitive detection of tumor biomarkers and cancer cells is of crucial importance for the early diagnosis and prognosis prediction of cancer. The present report describes a target-induced fluorescence enhancement immunosensor that utilizes the optical property of carbon dots (CDs) and the metal-enhanced fluorescence effect (MEF) property of silver nanoparticles (AgNPs) for the sensitive detection of the cancer biomarker human epididymis protein 4 (HE4) and ovarian cancer cells. Nitrogen and sulfur co-doped CDs with a quantum yield of 85.6% were prepared and served as the fluorophore in MEF. The HE4 antibody (Ab) specific to the HE4 antigen was linked covalently to the surface of the synthesized CDs as the capture. The HE4 Ab-conjugated AgNPs (AgNPs-Ab) were prepared and utilized as signal amplification elements. In the presence of the target HE4, composite sandwich structures were formed between the labeled CDs-Ab and AgNPs-Ab, which brought the CDs and AgNPs into proximity, resulting in the fluorescence of CDs enhancement owing to MEF. The intensity of fluorescence enhancement was positively correlated with the HE4 concentration in the clinically important range of 0.01–200 nM with a limit detection of 2.3 pM. Moreover, the immunosensor was also successfully applied to specific fluorescence labeling and quantitative determination of HE4-positive ovarian cancer cells. The proposed target-triggered MEF sensor platform demonstrated high sensitivity, excellent anti-interference ability, along with successful validation in complex biological matrices, providing a new approach for HE4 detection in early diagnosis and therapeutic monitoring.

Photothermal-Induced Electrochemical Interfacial Region Regulation Enables Signal Amplification for Dual-Mode Detection of Ovarian Cancer Biomarkers.

Detection sensitivity of an electrochemical immunosensor mainly depends on the accessible distance toward the sensing interface; regulating the electrochemical interfacial region thereon is an effective strategy for signal amplification. Herein, a photothermal-regulated sensing interface was designed based on a near-infrared (NIR)-responsive hydrogel probe for ultrasensitive detection of human epididymis protein 4 (HE4). Silver nanoparticle-deposited graphene oxide nanosheet (AgNPs@GO) hybrids as electrochemical signal tags and a photothermal transducer, which were encapsulated in the poly(N-isopropylacrylamide) (pNIPAM) hydrogel, were used to develop the NIR-responsive GO@AgNPs-pNIPAM hydrogel probe. Under NIR light irradiation, the excellent photothermal effect of AgNPs@GO hybrids not only rapidly converted NIR light into heat for temperature readout but also triggered the shrinkage behavior of the hydrogel for electrochemical signal amplification. Compared with the conventional sandwich immunoassay, the shrinkage behavior of the hydrogel signal probe endowed itself with interface regulation capability to improve the electrochemical reaction efficiency; on the basis of ensuring the extended outer Helmholtz plane (OHP) region, the proposed photothermal-induced interface regulation also shortened the OHP, leading to higher sensitivity. Moreover, the obtained dual-mode signals provided satisfactory accuracy for the detection of tumor markers. Therefore, this detection scheme provided an opportunity for the broad applications of the photothermal effect in clinical diagnosis.

Coupling metal-organic framework nanosphere and nanobody for boosted photoelectrochemical immunoassay of Human Epididymis Protein 4

A “signal-on” photoelectrochemical (PEC) immunosensor for highly sensitive detection of Human Epididymis Protein 4 (HE4), a new serum biomarker of ovarian cancer with small molecular weight, was fabricated by coupling the porphyrin-based metal-organic framework (MOF) nanosphere (nPCN-224) and Nanobody (Nb). To label the Nb, the nPCN-224 with an average size of 160–200 nm was prepared by solvothermal method. The mechanism for the photocurrent generation of nPCN-224 was systematically investigated, showing that the dissolved O2 in aqueous solution participated in the charge separation and transport during the photoelectric conversion by generating O2˙-, which resulted in a 6-fold enhanced photocurrent by using ascorbic acid as the O2 ˙- scavenger. Moreover, the inherent structural porosity of nPCN-224 demonstrated advantage for reactant accessibility. Due to the superior properties of nPCN-224, and the high specificity and affinity of Nbs, the immunosensor exhibited a broad detection range from 1.00 pg mL−1 to 10.0 ng mL−1 and a detection limit of 0.560 pg mL−1, lower than most methods reported before. The immunosensor could clearly distinguish ovarian cancer patients in different stages from healthy individuals, and the as-obtained results matched well with those by traditional electrochemiluminescence immunoassay method from the hospital. This work would open a new avenue for PEC immunosensors in clinical diagnostics and evaluation of potential clinical efficacy.

Diagnostic value of combined detection of carbohydrate antigen 125 and human epididymis protein 4 for epithelial ovarian cancer

Objective To explore the diagnostic value of combined detection of serum human epididymis protein 4 (HE4) and carbohydrate antigen 125 (CA125) for ovarian epithelial cancer. Methods Patients who underwent surgery for the adnexal tumor at Shanxi Provincial Cancer Hospital from January 2012 to December 2017 were enrolled. According to the postoperative pathological results, these patients were divided into the ovarian epithelial cancer group (494 cases) and benign ovarian disease group (462 cases). The serum expressions of HE4 and CA125 in the two groups were detected by enzyme-linked immunosorbent assay (ELISA) and chemiluminescence immunoassay. The diagnostic value of detection of HE4 and CA125 alone or in combination for ovarian epithelial cancer was analyzed. Results The median levels (P 25 - P 75) of serum CA125 and HE4 in ovarian epithelial cancer group were 273.34 U/ml (39.34 U/ml, 709.74 U/ml) and 199.08 pmol/L (75.81 pmol/L, 449.20 pmol/L), which were higher than those in ovarian benign disease group [16.30 U/ml (6.30 U/ml, 53.60 U/ml) and 39.54 pmol/L (29.57 pmol/L, 53.80 pmol/L)] (both P < 0.05). There was a positive correlation between serum CA125 and HE4 levels in ovarian epithelial cancer group (r = 0.481, P < 0.01). Serum CA125 and HE4 levels in patients with stage Ⅲ and Ⅳ were higher than those in patients with stage Ⅰ and Ⅱ (both P < 0.05), and serum CA125 and HE4 levels in patients with poor differentiation were higher than those in patients with moderate differentiation (P < 0.05). Compared with CA125, the specificity and positive predictive value of serum HE4 for the diagnosis of ovarian epithelial cancer were higher (both P < 0.01). Compared with HE4 alone, the sensitivity of CA125 combined with HE4 increased (P = 0.004) and the specificity decreased (P = 0.044). When both CA125 and HE4 were positive for positive results, compared with HE4 alone, the specificity and positive predictive value increased (both P < 0.01), but the sensitivity decreased (both P < 0.01). In patients with CA125+ HE4-, the sensitivity and specificity decreased, while in CA125- HE4+ patients, the specificity was elevated, but the difference was not statistically significant (P = 0.892). When one of CA125 and HE4 was positive for positive results, the sensitivity and negative predictive value increased (both P < 0.01), but the specificity and positive predictive value decreased (both P < 0.01). The area under the curve of serum CA125 and HE4 combined detection was 0.911, indicating its clinical diagnostic value was better than that of the two alone. Conclusions In the diagnosis of epithelial ovarian cancer, the specificity of serum HE4 is higher than that of CA125, but the sensitivity is lower than that of CA125. The combined detection of HE4 and CA125 is more conducive to improve the diagnostic accuracy of ovarian epithelial cancer. Key words: Ovarian neoplasms; Carbohydrate antigen 125; Human epididymis protein 4; Diagnosis

An ultrasensitive ratiometric electrochemiluminescence immunosensor combining photothermal amplification for ovarian cancer marker detection

Signal amplification strategies play important functions in the development of highly sensitive electrochemiluminescence (ECL) immunosensing system. Herein, a photothermal enhanced ratiometric ECL immunosensor was proposed for the detection of human epididymis protein 4 (HE4), an ovarian cancer biomarker. Mesoporous SiO2 (ms-SiO2) and carbon nanohorns (CNHs) was served as carrier of ECL emitter carbon nitride nanosheet (g-C3N4) and anodic ECL emitter polymer dots (Pdots), respectively. The large specific area of ms-SiO2 and CNHs improved the loading capacity of g-C3N4 and Pdots, enhancing ECL signals. Furthermore, CNHs was innovatively utilized as thermal convert unit to increase the electrode surface temperature, which benefited from its extraordinary photothermal property at 808 nm that can convert laser energy into heat for elevating the temperature, further amplified ECL signal. The delicately designed ECL immunosensor exhibited excellent sensitivity to HE4 detection with wide linear range from 1.0 × 10−5 to 10 ng/mL and low detection limit of 3.3 × 10−6 ng/mL. This work not only provided an effective way to develop highly sensitive ECL immunosensor but could attract more attention on the application of photothermal material in the ECL field.

Application of NiFe2O4 nanotubes as catalytically promoted sensing platform for ratiometric electrochemiluminescence analysis of ovarian cancer marker

Herein, a delicate ratiometric electrochemiluminescence biosensor was designed for the detection of human epididymis protein 4 (HE4) based on 2-(Dibutylamino)ethanol (DBAE) and lucigenin as anodic and cathodic ECL emitter, respectively. Initially, NiFe2O4 nanotubes (NiFe2O4 NTs) with charming properties could act as catalytically promoted sensing platform. This NiFe2O4 NTs based platform could trigger an intense ECL emission of DBAE for the reason that NiFe2O4 NTs not only achieved the high loading of Envision-Ab1 complex but effectively catalyzed the oxygen evolution reaction (OER) to yield oxygen. Moreover, the Envision complex, a polymeric material containing enrich horseradish peroxidase (HRP) could further accelerate the H2O2 decomposition to generate superoxide (O2−) for the enhancement of the ECL emission process of two luminophors. It’s noteworthy that hexagonal boron nitride nanosheets (h-BN) as the nanolabels, was first illustrated to be capable of immobilizing a large quantity of lucigenin and retaining intense and stable ECL emission of lucigenin in the alkaline condition. Benefiting from the above merits, a new ECL ratiometric sensing mode was established for the sensitive determination of HE4 in a range from 10 fg/mL to 10 ng/mL with the detection limit of 3.3 fg/mL. The strategy exhibited broad prospect in early ovarian cancer diagnosis and blazed a new path for other tumors screening.

Ultrasensitive Electrochemical Immunoassay Based on Cargo Release from Nanosized PbS Colloidosomes.

Colloidosome is a novel nanostructure composed of millions of colloid particles. In this work, nanosized PbS colloidosomes were initially prepared and applied as nanoprobes for an ultrasensitive immunoassay. The colloidosomes were simply prepared in mild conditions by assembling the elementary approximately 8 nm PbS nanoparticles at the water-in-oil interface of emulsion droplets. To enhance the rigidity and biocompatibility of the colloidosomes, interfacial polymer was introduced by utilizing self-polymerization of dopamine. By treating with dilute nitric acid, a bursting release of lead ions from the colloidosomes occurred and the lead ions can be detected easily by anodic stripping voltammetry. In this way, a colloidosome-based electrochemical immunoassay was developed by using the nanosized PbS colloidosomes as electroactive labels. The proposed method featured a linear calibration range from 10 fg·mL-1 to 100 ng·mL-1 with a low detection limit of 3.4 fg·mL-1 for the detection of human epididymis protein 4. This work introduced a new member for the family of colloidosomes and offered a novel perspective for the rational implementation of various colloidosomes for novel low-abundance cancer biomarkers analysis.

The diagnostic value of serum HE4 and CA-125 and ROMA index in ovarian cancer.

Ovarian cancer is a common malignancy of the female reproductive system. Tumor markers serve as tools in the diagnosis of the disease. The aim of the present study was to determine the diagnostic value of sera levels of carbohydrate antigen-125 (CA-125), human epididymis protein 4 (HE4) as well as the area under the curve of the receiver operating characteristic (ROC) and the risk of ovarian malignancy algorithm (ROMA) index in ovarian cancer. The sera were measured using an electrochemiluminescence immunoassay on 158 individuals (64 patients with ovarian cancer, 64 with ovarian benign tumor and 30 healthy individuals) between September 2013 and May 2015. The results showed that levels of HE4 and CA-125 in the sera of the ovarian benign tumor group as well as their ROMA index were significantly higher (P<0.05) than those of the ovarian benign tumor and control groups, regardless of pre- or postmenopausal status. However, the level of CA-125 was significantly higher (P<0.05) in the ovarian benign tumor group compared with the healthy group, while the level of HE4 was similar in the two groups. The sensitivity of the ROMA index was higher (P<0.01) with detection of HE4 and CA-125. In the ovarian cancer group, the areas under ROC curves of ROMA, HE4 and CA-125 were 0.994, 0.990 and 0.941, respectively. The specificity and positive predictive value of HE4 in the premenopausal ovarian cancer group reached 98.36 and 95%, respectively. In conclusion, the results showed that the serum level of HE4 and the ROMA index are important indicators in the diagnosis of ovarian cancer. However, in addition to HE4 and CA-125 detection, the ROMA index is extremely valuable in improving the diagnostic efficiency of ovarian cancer.

The development of dual-label time-resolved fluorescence immunoassay (TRFIA) for screening of ovarian cancer based on simultaneous detection of human epididymis protein-4 and cancer antigen 125

ABSTRACTA two-step dual-label TRFIA was developed for the simultaneous detection of human epididymis protein-4 and cancer antigen 125 in a single run. The performance of this assay was first evaluated using clinical serum samples, and then compared with commercialized kits. The sensitivity of this assay for cancer antigen 125 detection was 0.5 U/mL (dynamic range, 0–1400 U/L), and the sensitivity for human epididymis protein-4 detection was 1 pM (dynamic range, 1–900 pM). High correlation coefficients (R) were obtained between the present dual-label TRFIA and commercially available kits (R = 0.99). The present dual-label TRFIA has high sensitivity, specificity, and accuracy in clinical sample analysis. It is a good alternative to the single-label diagnostic methods.

The diagnosis and pathological value of combined detection of HE4 and CA125 for patients with ovarian cancer.

ObjectiveTo evaluate the value of individual and combined measurement of human epididymis protein 4 (HE4) and cancer antigen 125 (CA-125) in the diagnosis of ovarian cancer.MethodsA clinical case-control study was performed in which the levels of serum HE4 and CA-125 of subjects with malignant, borderline, benign ovarian tumors and healthy women were measured before surgery. An immunohistochemistry method was used to measure the expression of HE4 in different tissues. Statistical analysis was performed to determine the relationship between the level of HE4 and the pathologic type as well as the stage of the ovarian tumors.ResultsThe level of HE4 in the serum was significantly elevated in the malignant ovarian cancer group compared with other groups. Women with benign ovarian tumors and non-neoplastic lesions, and healthy women were designated as references. When the level of HE4 in the serum was 58.66 pmol/L, the sensitivity and specificity of HE4 in diagnosing malignant ovarian tumors was 82.35% and 96.03%, respectively. The level of HE4 was negatively correlated with the differentiation extent of the tumors whereas positively correlated to the clinical staging. In the groups of malignant and borderline tumors, the levels of HE4 were higher than the other groups. The expression of HE4 was significant higher in the serous types of ovarian tumors than that of the mucous types (P<0.05). The level of HE4 in the serum and tissues were positively correlated with each other.ConclusionHE4 can be used as a novel clinical biomarker for predicting malignant ovarian tumors and its expression was closely related with the clinical pathological features of malignant ovarian tumors.

Detection of Human Epididymis Protein 4 (HE4) in Human Serum Samples Using a Specific Monoclonal Antibody-Based Sandwich Enzyme-Linked Immunosorbent Assay (ELISA).

The human epididymis protein 4 (HE4) may have high specificity in the detection of malignant diseases, making the development of an immunoassay for HE4 essential. In our study, a fusion gene was constructed encoded with the HE4 protein. This protein was then produced in the bacterial cells (Escherichia coli) and used to immunize mice in order to eventually generate hybridomas specific to HE4. The hybridoma supernatants were then screened, and four positive anti-HE4 cell lines were selected. These cell lines produce monoclonal antibodies against HE4 epitopes, as demonstrated in the Western blot as well as by direct enzyme-linked immunosorbent assay (ELISA). Using the developed antibodies, we successfully identified several good antibody pairs from the hybridomas, which allowed for the development of a sandwich ELISA to measure HE4 levels. By using the HE4 ELISA, we measured HE4 levels of 60 clinical human serum samples. Compared with the Food and Drug Administration (FDA) approved kit (Roche), our results showed a strong positive correlation to those of the FDA-approved kit. In summary, highly sensitive antibody pairs were screened against HE4, and a sandwich ELISA was developed as an accurate analytical tool for the detection of HE4 in human serum, which could be especially valuable for diagnosing ovarian carcinomas.

Alkaline phosphatase labeled antibody-based electrochemical biosensor for sensitive HE4 tumor marker detection

Quantitative determination of the serum level of human epididymis protein 4 (HE4), a tumor marker for ovarian carcinoma, has come to the fore of interest mainly due to the possibility for its detection in early stages of the disease when the sensing of other biomarkers is limited. We present a simple ELISA based approach for rapid HE4 detection including its immunomagnetic capturing accompanied by sensitive electrochemical detection of the electroactive product formed after enzymatic conversion of the substrate by alkaline phosphatase used as a label of anti-HE4 IgG. The proposed immunosensor offers stability in time and due to excellent limit of detection at 6.8 fM HE4, and limit of quantification of 23 fM meets the requirements for the early detection of this biomarker.

Investigation on the application significance of serum tumor biomarker HE4 detection in the diagnosis of lung cancer

Objective To investigate the expression level of tumor biomarker human epididymis protein 4( HE4)in tissue and serum of patients with lung cancer and its role in differential diagnosis of lung cancer. Methods HE4 expressions in lung adenocarcinoma,lung squamous cell carcinoma and cancer-adjacent tissues were detected by immunohistochemistry,and serum HE4 levels were determined in 214 patients with lung cancer and 76 healthy subjects by enzyme-linked immunobsorbent assay( ELISA). Simultaneously,the serum levels of squamous cell carcinoma antigen( SCC),carcinoembryonic antigen( CEA),cytokeratin 19 fragment( CYFRA21-1) and neuron-specific enolase( NSE) were determined. The results were compared and analyzed among the groups. Results Lung adenocarcinoma tissues showed stronger positive staining than lung squamous cell carcinoma tissues,and the negative staining was shown in cancer-adjacent tissues. The level of serum HE4 in lung cancer patients[85. 04( 64. 91,118. 40) pmol /L] was significantly higher than that of healthy controls [48. 48( 43. 12,58. 81) pmol /L,P 0. 01]. HE4 had a better diagnosis efficiency than CYFRA21-1 and CEA in non-small cell lung cancer through receiver operating characteristic( ROC) curve analysis. The positive rate of combined detection of SCC,CEA,CYFRA21-1 and NSE was 79.44% for lung cancer. However,HE4 improved the positive rate to 89. 72% when combined with this biomarker panel. In the diagnosis of lung adenocarcinoma,the positive rate of combined detection of CEA and CYFRA21-1 was 68. 46%.Moreover,HE4 improved the positive rate to 82. 31% when combined with this biomarker panel. Conclusions HE4,as a biomarker of lung cancer,is a useful indicator for the early diagnosis of lung cancer.

HE4 as a Serum Biomarker for ROMA Prediction and Prognosis of Epithelial Ovarian Cancer

Human epididymis protein 4 (HE4) has been suggested to be a novel biomarker of epithelial ovarian cancer (EOC). The present study aimed to evaluate and compare HE4 with the commonly used marker, carbohydrate antigen 125 (CA125), in prediction and therapy-monitoring of EOC. Serum HE4 concentrations from 123 ovarian cancer patients and 174 controls were measured by Roche electrochemiluminescent immunoassay (ECLIA). Risk of ovarian malignancy algorithm (ROMA) values were calculated and assessed. In addition, the prospects of HE4 detection for therapy-monitoring were evaluated in EOC patients. The ROMA score could classify patients into high- and low-risk groups with malignancy. Indeed, lower serum HE4 was significantly associated with successful surgical therapy. Specifically, 38 patients with EOC exhibited a greater decline of HE4 compared with CA125. In contrast, elevation of HE4 better predicted recurrence (of 46, 11 patients developed recurrence, and with it increased HE4 serum concentrations) and a poor prognosis than CA125. This study suggests that serum HE4 levels are closely associated with outcome of surgical therapy and disease prognosis in Chinese EOC patients.