AlphaLISA-Based Immunoassay for Detection of Troponin T in Serum of Patients with Acute Myocardial Infarction.

Survival and prognosis of patients with acute myocardial infarction (AMI) are highly dependent on rapid and accurate diagnosis of myocardial damage. Troponin T is the primary diagnostic biomarker and is widely used in clinical practice. Amplified luminescent proximity homogeneous assay (AlphaLISA) may provide a solution to rapidly detect a small amount of analyte through molecular interactions between special luminescent donor beads and acceptor bead. Here, a double-antibody sandwich assay was introduced into AlphaLISA for rapid detection for early diagnosis of AMI and disease staging evaluation. The performance of the assay was evaluated. The study found that the cTnT assay has a linear range of 48.66 to 20,000 ng/L with a limit of detection of 48.66 ng/L. In addition, the assay showed no cross-reactivity with other classic biomarkers of myocardial infarction and was highly reproducible with intra- and inter-batch coefficients of variation of less than 10%, notably, only 3 min was taken, which is particularly suitable for clinical diagnosis. These results suggest that our method can be conveniently applied in the clinic to determine the severity of the patient's condition.

Establishment of trypsinogen-2 Amplification Luminescent Proximity Homogeneous Assay and its Application in Acute Pancreatitis.

We aim to develop an amplified luminescence proximity homogeneous assay (AlphaLISA) for quantification of trypsinogen-2 levels in human serum for the diagnosis of acute pancreatitis. Based on new amplified luminescence proximity homogeneity assay (AlphaLISA) method, carboxyl-modified donor and acceptor beads were coupled to capture and detection antibodies. A double antibody sandwich immunoassay was used to detect the concentration of trypsinogen-2 in serum. The method had good linearity (> 0.998). The intra - analysis precision was between 1.54% and 2.20% (< 10%), the inter-analysis precision was between 3.17% and 6.94% (< 15%), and the recovery was between 96.23% and 103.45%. The cross-reactivity of carbohydrate antigen 242 (CA242) and T-cell immunoglobulin mucin-3 (Tim-3) were 0.09% and 0.93%, respectively. The detection time only needed 15 min. The results of trypsinogen-2-AlphaLISA and time-resolved fluorescence immunoassay were consistent (ρ = 0.9019). In addition, serum trypsinogen-2 concentration in patients with acute pancreatitis [239.23 (17.83-807.58) ng/mL] was significantly higher than that in healthy controls [20.54 (12.10-39.73) ng/mL]. When the cut-off value was 35.38ng/mL, the sensitivity and specificity were 91.8% and 96.67%, and the positive detection rate was 91.80%. We have successfully established a trypsinogen-2-AlphaLISA method, which can promote the timely diagnosis of acute pancreatitis.

Advances in Research on the Regulation of Floral Development by CYC-like Genes.

CYCLOIDEA (CYC)-like genes belong to the TCP transcription factor family and play important roles associated with flower development. The CYC-like genes in the CYC1, CYC2, and CYC3 clades resulted from gene duplication events. The CYC2 clade includes the largest number of members that are crucial regulators of floral symmetry. To date, studies on CYC-like genes have mainly focused on plants with actinomorphic and zygomorphic flowers, including Fabaceae, Asteraceae, Scrophulariaceae, and Gesneriaceae species and the effects of CYC-like gene duplication events and diverse spatiotemporal expression patterns on flower development. The CYC-like genes generally affect petal morphological characteristics and stamen development, as well as stem and leaf growth, flower differentiation and development, and branching in most angiosperms. As the relevant research scope has expanded, studies have increasingly focused on the molecular mechanisms regulating CYC-like genes with different functions related to flower development and the phylogenetic relationships among these genes. We summarize the status of research on the CYC-like genes in angiosperms, such as the limited research conducted on CYC1 and CYC3 clade members, the necessity to functionally characterize the CYC-like genes in more plant groups, the need for investigation of the regulatory elements upstream of CYC-like genes, and exploration of the phylogenetic relationships and expression of CYC-like genes with new techniques and methods. This review provides theoretical guidance and ideas for future research on CYC-like genes.

Establishment of 11-dehydro-thromboxane B2 time-resolved immunoassay and application in membranous nephropathy.

BACKGROUND: 11-Dehydro-thromboxane B2 (11-dehydro-TXB2) is the final stable metabolite of thromboxane A2 (TXA2) and is involved in thrombus formation. Patients with membranous nephropathy (MN) are prone to thromboembolism events. METHODS: Time-resolved fluorescence immunoassay (TRFIA) for 11-dehydro-TXB2 was established by indirect competitive method. The coated 11-dehydro-TXB2-BSA conjugate was used to bind the 11-dehydro-TXB2 antibody competitively to the 11-dehydro-TXB2 antigen in the samples, followed by Eu3+-labeled goat anti-mouse IgG antibody, to detect 11-dehydro-TXB2. This study measured 11-dehydro-TXB2 concentrations in serum samples from healthy individuals and patients with MN. RESULTS: The linear range of TRFIA was 16.38-2000 pg/mL, the sensitivity was 4.70 pg/mL, the average coefficients of variation from intra-assay and inter-assay were 3.50% and 4.95%, respectively, and the recovery was 99.38%. The serum level of 11-dehydro-TXB2 in patients with MN was significantly higher than that in healthy subjects (P < 0.05). The serum 11-dehydro-TXB2 concentration detected by TRFIA was highly consistent with that by ELISA (ρ = 0.900). DISCUSSION: This study successfully established a new highly sensitive method for the detection of 11-dehydro-TXB2 in serum. 11-Dehydro-TXB2 has great potential in evaluating the risk of thromboembolic events in patients with MN and is expected to be applied to other thromboembolic-related diseases.

Establishment of matrix metalloproteinase 3 time-resolved immunoassay and some potential clinical applications.

AIM: To develop a highly sensitive time-resolved fluorescence immunoassay (TRFIA) for the detection of serum matrix metalloproteinase-3 (MMP-3) and to assess MMP-3's clinical value in patients with colorectal cancer (CRC).st. METHODS: MMP-3 levels were established using the double antibody sandwich technique. The MMP-3 TRFIA technique was developed and optimized, and its linearity, sensitivity, accuracy, specificity, and recovery were assessed. Then, serum concentrations in healthy individuals and patients with CRC were determined by MMP-3 TRFIA. RESULTS: The linear range of MMP-3 TRFIA was 0.73-500 ng/mL. MMP-3 TRFIA had an intra-batch precision range of 2.16%-7.10% percent and an inter-batch precision range of 3.99%-11.21%. MMP-3, tumor-associated trypsinogen 2, and AFP had no cross reaction.The recovery is between 90% and 110%, and had no serum interference. Patients with CRC had serum MMP-3 levels (73.95 ± 78.43 ng/mL) that were considerably higher than those of healthy individuals (21.45 ± 11.12 ng/mL), and those with metastasis had serum MMP-3 levels (95.89 ± 76.21 ng/mL) that were considerably higher than those of patients without metastasis (52.74 ± 47.25 ng/mL). CONCLUSIONS: A highly sensitive MMP-3 TRFIA assay was successfully developed, and serum MMP-3 may be associated with CRC invasion and metastasis. Therefore, MMP-3 can be used in the auxiliary diagnosis of CRC.

Development of amplified luminescent proximity homogeneous assay for quantitation of gastrin-17.

A highly sensitive and convenient amplified luminescent proximity homogeneous assay (AlphaLISA) method with high throughput and automation potential was developed for quantitation of serum Gastrin-17 (G-17) levels, which can facilitate the early diagnosis of atrophic gastritis in people at high risk of gastric cancer using a non-invasive approach. In this study, donor and acceptor beads with modified carboxyl groups on the surface were directly coupled to anti-G-17 antibodies through activation was proposed for application in the development of the new AlphaLISA, which can effectively simplify the steps and shorten the reaction time to achieve faster detection. Therefore, the G-17-AlphaLISA only needs to react for 15 min to obtain good analysis results. The proposed method has a wider detection range than commercial enzyme-linked immunosorbent assay (ELISA) kits (0.12-112.8 pmol/L > 0.5-40 pmol/L). In addition, results of G-17-AlphaLISA and ELISA had good correlation and agreement (ρ = 0.936). Importantly, the developed method may be more suitable for the large-scale screening of people at high risk for gastric cancer than traditional ELISA and provides a novel solution for other biomarkers that require accurate, highly sensitive, and high throughput detection.

Quantitative detection of anti-PLA2R antibodies targeting different epitopes and its clinical application in primary membranous nephropathy.

OBJECTIVES: This study aimed to establish time-resolved fluorescence immunoassays to quantitatively detect the autoantibodies targeting different epitopes of M-type phospholipase A2 receptor (PLA2R) and evaluate its clinical application in primary membranous nephropathy (PMN). METHODS: PLA2R and its reactive epitope-specific IgG/IgG4 time-resolved fluorescence immunoassays (TRFIAs) were established using europium-labeled anti-human IgG/IgG4 antibodies, recombinant proteins, and patient serum. The levels of IgG/IgG4 targeting PLA2R and its epitopes in PMN patient serum were detected, and the relationship between epitope spreading of PLA2R and the severity of patients with PMN was evaluated. RESULTS: The TRFIAs established in this study could quantitatively detect PLA2R and its epitope-specific IgG and IgG4. Sera from 59 patients with PMN were subjected to detection using anti-PLA2R IgG and anti-PLA2R IgG4. Among them, 46 and 54 patients were found positive for PLA2R antibodies, respectively. Moreover, the levels of PLA2R antibodies were strongly correlated with the severity of patients with PMN. Patients who were detected to have two or more epitopes had more serious renal injury. CONCLUSIONS: PLA2R domain-specific IgG/IgG4 TRFIAs were established in this study, and detection with anti-PLA2R IgG4 could more sensitively screen the reactivity of patients to the PLA2R domain. Moreover, detection epitope spreading of PLA2R was confirmed which is related to the severity of patients with PMN.

Establishment and Clinical Application in Stroke of a Serum Copeptin Time-Resolved Fluorescence Immunoassay.

The serum biomarker copeptin, an innovative and stable substitute biomarker of vasopressin, is associated with stroke. Therefore, establishing a highly sensitive time-resolved fluorescence immunoassay for copeptin (copeptin-TRFIA) is helpful to measure stroke and evaluate its value in clinical applications. Double antibody sandwich was used to establish copeptin-TRFIA. The established method was then assessed. Two coated and Eu3+-labeled copeptin monoclonal specific antibodies targeting different antigen epitopes were employed. The serum fluorescence counts of patients with stroke and healthy volunteers were detected by using the well-established copeptin-TRFIA. Serum copeptin levels were measured and analyzed statistically. The actual measurement linearity range of the proposed method was 0.13-44.66 ng/mL. Copeptin-TRFIA had the inter-assay coefficient of variation (CV) of 6.49%-9.08% and the intra-assay CV of 4.75%-7.77%. Patients with cerebral infarction (CI) and intracerebral hemorrhage (ICH) had significantly higher serum copeptin levels than healthy subjects. Copeptin concentrations in the serum of patients with stroke were significantly correlated with the scores of the National Institute for Healthy Stroke Scale (NIHSS) and modified Rankin Scale (mRS). A highly sensitive copeptin-TRFIA was successfully established. Serum copeptin has a certain value in the clinical diagnosis and prognosis of stroke.

Establishment and clinical application of time-resolved immunofluorescence assay of lipoprotein-associated phospholipase A2.

AIM: This study aimed to establish a highly sensitive time-resolved fluorescence immunoassay (TRFIA) for the detection of serum lipoprotein-associated phospholipase A2 (Lp-PLA2) and evaluate the clinical application value of Lp-PLA2 in patients with breast cancer. METHODS: The level of Lp-PLA2 was detected using the double-antibody sandwich method. First, the Lp-PLA2-TRFIA method was established, and the method was evaluated on the basis of linearity, sensitivity, precision, specificity, and recovery rate. Then, the fluorescence counts in serum of healthy subjects and patients with breast cancer were detected by Lp-PLA2-TRFIA, and the levels of Lp-PLA2 were calculated using a standard curve. RESULTS: Lp-PLA2-TRFIA had a wide linear range (43.48-2000 ng/mL). The intra-assay precisions of Lp-PLA2-TRFIA ranged from 2.66% to 4.84% (<10%), and the inter-assay precisions were between 5.39% and 6.95% (<15%). No cross-reaction was observed among Lp-PLA2, Tumor-associated trypsinogen-2, and T-cell immunoglobulin mucin 3. In addition, the recovery rates were between 90% and 100%. The serum Lp-PLA2 levels of patients with breast cancer were significantly higher than those of healthy subjects. CONCLUSIONS: We successfully established a highly sensitive Lp-PLA2-TRFIA method, and found serum Lp-PLA2 may be associated with dyslipidemia in breast cancer and could be used for auxiliary diagnose.

Development and application of amplified luminescent proximity homogeneous assay for quantitation of heparin-binding protein.

A fast and highly sensitive amplified luminescent proximity homogeneous assay (AlphaLISA) method was developed for quantitation of plasma heparin-binding protein levels. In this study, a method directly coupling donor and acceptor beads modified with aldehyde groups to anti-HBP antibodies was proposed, which can effectively simplify the steps and shorten the reaction time to achieve faster detection. Therefore, the developed method required only 15 min of reaction time to generate results. Compared with the approved commercial kit, the developed method had a wider linear range (2.78-500 ng/mL). The excellent linear range means that the method can better exploit the value of HBP in clinical applications. Meanwhile, results of amplified luminescent proximity homogeneous assay and fluorescence dry quantitative immunoassay had good correlation and consistency (ρ = 0.9181). Moreover, the plasma HBP concentrations of patients with bacterial infection were significantly higher than those of healthy individuals (P < 0.0001), indicating the potential applicability of the proposed method for predicting the incidence of bacterial infections. Importantly, the newly developed method is expected to serve as an alternative to the traditional assay method and provides a completely new platform for other biomarkers that require rapid detection.

Establishment and Clinical Application of a Highly Sensitive Time-Resolved Fluorescence Immunoassay for Tumor-Associated Trypsinogen-2.

To establish a rapid and highly sensitive assay for tumor-associated trypsinogen-2 (TAT-2) based on the time-resolved fluorescence immunoassay (TRFIA) and evaluate its potential clinical value in patients with lung cancer. The double-antibody sandwich method was used in detecting TAT-2 antigen concentrations, and two types of TAT-2 antibodies (coating antibodies and Eu3+ labeled antibodies) were used. A TAT-2-TRFIA method was then established, evaluated, and used in detecting the serum TAT-2 levels of healthy subjects and patients with lung cancer. The linear range of the TAT-2-TRFIA method was 1.53-300 ng/mL, the intra-assay coefficient of variation (CV) were between 1.67% and 8.42%, and the inter-assay CV were between 4.29% and 11.44%. The recovery rates of TAT-2-TRFIA were between 99.17% and 107.06%. The cross-reactivities of trypsin and T-cell immunoglobulin mucin 3 were 0.02% and 0.82%, respectively. The serum TAT-2 levels of patients with lung cancer were higher than those of healthy subjects (P < 0.001). Combined with TAT-2, the sensitivity and specificity of CEA and CA-125 for lung cancer improved significantly. Conclusion: We successfully established a highly sensitive TAT-2-TRFIA method, which was able to facilitate the timely diagnosis of lung cancer.

Establishment of Galectin-3 Time-resolved Fluoroimmunoassay and its Application in Idiopathic Membranous Nephropathy.

The aim of this study was to establish a time-resolved fluorescent immunoassay (TRFIA) for the detection of serum Galectin-3 (Gal-3) and apply this method to evaluate the clinical significance of serum Gal-3 in predicting Idiopathic Membranous Nephropathy (IMN) progression. The Gal-3-TRFIA was established using the double antibody sandwich method, with the capture antibodies coated on a 96-well microplate and the detection antibodies chelated with Europium (III) (Eu3+). Serum Gal-3 was detected in 81 patients with IMN and 123 healthy controls to further evaluate the value of the Gal-3 in staging of IMN. The sensitivity of the Gal-3-TRFIA assay was 0.85 ng/mL, and the detection range was 0.85-1000 ng/mL. The Gal-3 intra-batch and inter-batch coefficients of variation were 3.45% and 5.12%, respectively. The correlation coefficient (R) between the Gal-3-TRFIA assay and commercially available enzyme-linked immunosorbent assay kits was 0.83. The serum Gal-3 concentration was higher in patients with IMN (65.57 ± 55.90 ng/mL) compared to healthy controls (16.29 ± 9.91 ng/mL, P < 0.0001). In this study, a wide detection range Gal-3-TRFIA assay was developed using lanthanide (Eu3+) chelates for the detection of Gal-3 concentrations in serum. Gal-3 concentration is elevated in patients with IMN.

Establishment of a time-resolved immunoassay for acute kidney injury based on the detection of Kim-1.

AIM: To establish a highly sensitive time-resolved fluorescence immunoassay (TRFIA) of kidney injury molecule-1 (Kim-1) and evaluate its clinical value in acute kidney injury (AKI). METHODS: The Kim-1-TRFIA was established by the double-antibody sandwich method, and the method was evaluated. The established Kim-1-TRFIA was used to detect the concentration of Kim-1 in the serum of healthy controls and patients with AKI. RESULTS: The optimal coating antibody concentration and optimal Eu3+ -labeled antibody dilution ratio for Kim-1-TRFIA are 1 µg/ml and 1:140, respectively. The linear range is 42.71-4666.69 pg/ml. The intra- and inter-assay coefficients of variation are <10%. The specificity of our Kim-1-TRFIA is acceptable. The recovery is between 95.14% and 102.84%. The concentration of Kim-1 in the serum of patients with AKI is 126.50 ± 67.99 pg/ml, which is significantly higher than that in the serum of healthy controls (49.72 ± 16.40 pg/ml, p < 0.001). Staging patients with AKI by glomerular filtration rate shows that the serum concentration of Kim-1 increases significantly with increasing disease severity (p < 0.05). CONCLUSION: A highly sensitive Kim-1-TRFIA was established. With this immunoassay, a good differential diagnosis can be made, and healthy people and AKI patients can be differentiated by detecting the concentration of Kim-1 in the serum. Moreover, the severity of AKI patients can be determined.

Establishment of heparin-binding protein time-resolved immunoassay and some potential clinical applications.

AIM: To establish a highly sensitive time-resolved fluorescence immunoassay of heparin-binding protein (HBP-TRFIA) and evaluate its application value for bacterial or fungal infections in tumor patients. METHODS: Two types of HBP monoclonal specific antibodies against different epitopes of the antigen molecule were used as coating antibodies and Eu3+-labeled antibodies, respectively. The double-antibody sandwich method was used in establishing HBP-TRFIA, and the methodology was evaluated. The established HBP-TRFIA was used in detecting HBP concentration in the plasma samples of healthy individuals, patients with bacterial or fungal infections, and infected or uninfected patients with various types of tumors. RESULTS: The linear range of HBP-TRFIA was (0.11-530 ng/mL). Plasma HBP concentrations detected through HBP-TRFIA were consistent with the results of fluorescence quantitative immunochromatography (ρ = 0.964). The plasma HBP concentrations of infected tumor patients were significantly higher than those of uninfected tumor patients (P < 0.01). CONCLUSION: This study successfully established a highly sensitive HBP-TRFIA, which was highly comparable to commercially available fluorescent quantitative immunochromatographic kits and was able to facilitate the timely diagnosis of bacterial or fungal infections in patients with tumor.

Establishment of time-resolved fluoroimmunoassay of IgG4 based on magnetic microspheres.

BACKGROUND: The abnormal increase in serum IgG4 level is an important clinical symptom of IgG4-related disease (IgG4-RD), and the detection of serum IgG4 level is a powerful tool for the diagnosis of IgG4-RD. This study was conducted to establish a simple and rapid immunoassay for the determination of human serum IgG4 levels. METHODS: Based on the competition method, a novel immunoassay was established for the determination of human serum IgG4 using a combination of time-resolved fluoroimmunoassay (TRFIA) and magnetic microspheres. IgG4 was coupled with magnetic microspheres and competed with IgG4 in the samples to bind the Eu3+ -labeled anti-IgG4 antibody. The immunocomplex was separated and washed in a magnetic field, and the fluorescence counts were measured according to the number of dissociated europium ions. RESULTS: The analytical sensitivity of IgG4-TRFIA based on magnetic microspheres was 0.006 g/L, and the detection range was 0.006-20 g/L under optimal conditions. The precision, recovery, and specificity of this immunoassay were demonstrated to be acceptable. The clinical application of IgG4-TRFIA based on magnetic microspheres was evaluated and compared with that of immunonephelometry. The results showed that the two detection methods had a good correlation, with a correlation coefficient of .9871. CONCLUSION: IgG4-TRFIA based on magnetic microspheres has the advantages of high sensitivity, wide detection range, and short analysis time and has the potential to become a useful tool for the diagnosis of IgG4-RD.

Establishment of interleukin-18 time-resolved fluorescence immunoassay and its preliminary application in liver disease.

BACKGROUND: To establish a time-resolved fluorescence immunoassay of interleukin (IL)-18 (IL-18-TRFIA) and detect its concentration in different liver disease serum samples. METHODS: The IL-18 coating antibody and the Eu3+ -labeled detection antibody were used for the IL-18-TRFIA to detect serum IL-18 concentration in patients with liver cancer, hepatitis B, hepatitis C, autoimmune hepatitis, fatty liver disease, and healthy controls. The double-antibody sandwich method was used and methodological evaluation was performed. RESULTS: The average intra- and inter-assay coefficient of variation for IL-18-TRFIA was 4.80% and 5.90%, respectively. The average recovery rate was 106.19 ± 3.44%. The sensitivity (10.96 pg/mL) was higher than that obtained using the ELISA method (62.5 pg/mL). The detection range was 10.96-1000 pg/mL. IL-6 and galectin-3 did not cross-react with IL-18-TRFIA. The serum concentration of IL-18 was (776.99; 653.48-952.39 pg/mL) in hepatitis C, (911; 775.55-1130.03 pg/mL) in fatty liver, (1048.88; 730.04-1185.10 pg/mL) in liver cancer, and (949.12; 723.70-1160.28 pg/mL) in hepatitis B. Moreover, IL-18 serum levels were significantly higher in patients than the healthy controls (483.09; 402.52-599.70/mL) (p < 0.0001). Autoimmune hepatitis with a serum IL-18 concentration of 571.62; 502.47-730.31 pg/mL was not significantly different from the healthy controls (p > 0.05). CONCLUSION: We established a highly sensitive IL-18-TRFIA method that successfully detected serum IL-18 concentrations in different liver diseases. Furthermore, IL-18 serum concentration was higher in patients with liver cancer, hepatitis C, hepatitis B, and fatty liver disease compared to healthy controls.

Doxorubicin hydrochloride enhanced antitumour effect of CEA-regulated oncolytic virotherapy in live cancer cells and a mouse model.

Oncolytic adenovirus (OA) has attracted increasing attention due to their specific proliferation in tumour cells and resulting in lysis of tumour cells. To further improve the antitumour effect of OA, in this study, we combined CD55-TRAIL-IETD-MnSOD (CD55-TMn), a CEA-controlled OA constructed previously, and chemotherapy to investigate their synergistic effect and possible mechanisms. MTT assay was performed to detect antitumour effects. Hoechst 33 342 and flow cytometric analysis were used to examine cell apoptosis. Western blotting was performed to examine cell pyroptosis and apoptosis mechanism. Animal experiment was used to detect antitumour effect of doxorubicin hydrochloride (Dox) combined with CD55-TMn in vivo. We firstly found that Dox promotes gene expression mediated by CEA-regulated OA and virus progeny replication by activating phosphorylation of Smad3, and Dox can enhance antitumour effect of CEA-regulated CD55-TMn by promoting cell apotopsis and cell pyroptosis. Thus, our results provide an experimental and theoretical basis on tumour therapy by combination treatment of the oncolytic virotherapy and chemotherapy and it is expected to become a novel strategy for liver cancer therapy.

Soluble Tim3 detection by time-resolved fluorescence immunoassay and its application in membranous nephropathy.

BACKGROUND: We aimed to develop a time-resolved fluorescence immunoassay (TRFIA) for detecting soluble T-cell immunoglobulin and mucin domain 3 (sTim3) in serum samples and to demonstrate a preliminary application of this method in membranous nephropathy (MN). METHODS: sTim3 TRFIA was developed, and the sTim3 concentration in the serum of patients with MN and healthy individuals was detected using a sandwich method. RESULTS: The sensitivity of the developed sTim3 TRFIA was 0.66 ng/mL, higher than that of an enzyme-linked immunosorbent assay (ELISA) (1.11 ng/mL). The detection range was 0.66-40 ng/mL. The intra-assay coefficient of variation (CV) for sTim3 was 1.64%-4.68%, and the inter-assay CV was 5.72%-9.32%. The cross-reactivity to interleukin 6 (IL-6) and kidney injury molecule 1 (KIM-1) was 0.25% and 0.04%, respectively. The average recovery was 105.26%. The sTim3 concentration in patients with MN was considerably higher than that in healthy individuals (P < .001). The sTim3 concentration in the serum of patients with MN was significantly increased from G1 to G4 based on the Jonckheere-Terpstra test (P < .001). Thus, we used sTim3 as a diagnostic indicator for distinguishing between healthy individuals and patients with MN as well as between different stages of MN. CONCLUSION: We successfully established TRFIA to detect sTim3 in serum. We then applied this method to patients with MN, demonstrating for the first time that TRFIA is a valid diagnostic tool to detect sTim3 in serum.

Association between nucleotide excision repair gene polymorphism and colorectal cancer risk.

BACKGROUND: The nucleotide excision repair system removes a wide variety of DNA lesions from the human genome, and plays an important role in maintaining genomic stability. Single nucleotide polymorphisms (SNPs) in nucleotide excision repair are associated with the various forms of tumor susceptibility. However, the relationship between NER polymorphism and colorectal cancer is not clear. METHODS: In this study, three candidate SNPs including ERCC4 (rs6498486), ERCC1 (rs3212986), and ERCC5 (rs17655) were analyzed in 1101colorectal cancer patients and 1175 healthy control patients from Jiangsu province (China). Then, we performed Immunohistochemistry, qPCR, and luciferase assay to determine the potential mechanisms. RESULTS: The ERCC4 rs6498486 AC/CC genotypes show lower susceptibility to CRC than those carrying rs6498486 AA (Adjusted OR = 0.82, 95% CI = 0.69-0.97). However, we did not observe any association between the colorectal cancer risk and the rs3212986(ERCC1) and rs17655(ERCC5) polymorphisms. Immunohistochemistry, qPCR, and luciferase assay revealed that rs6498486 A > C polymorphism in the ERCC4 promoter region could lessen the expression level of ERCC4 by impacting the binding ability of the transcription factor NF-kB, thereby affecting the transcription activity of the ERCC4 gene and decreased ERCC4 gene expression. CONCLUSION: In brief, our finding demonstrated that ERCC4 rs6498486 serves as a potential biomarker of CRC susceptibility for the development of colorectal cancer.

GOLPH2-regulated oncolytic adenovirus, GD55, exerts strong killing effect on human prostate cancer stem-like cells in vitro and in vivo.

GOLPH2 (also called GP73) is a Golgi glycoprotein, which has been identified as a novel tumor marker upregulated in various cancers, including prostate cancer (PCa). GD55 is a novel GOLPH2-regulated oncolytic adenovirus that exhibits a strong killing effect on hepatoma cells. Here, we investigate the antitumor effect of GD55 on prostate cancer stem cell (CSC)-like cells in vitro and in vivo. Prostate CSC-like sphere cells were acquired and enriched by culturing DU145, LNCap or P3 prostate cancer cells in suspension. The prostate CSC-like sphere cells were capable of self-renewal, differentiation and quiescence, displaying tumorigenic feature and chemo-resistance to 5-FU, doxorubicin and DDP. Treatment with GD55 (1, 5, 10 MOI) dose-dependently suppressed the viability of DU145 sphere cells, which was a more pronounced compared to its cytotoxic action on the parental DU145 cells. In a mouse xenograft prostate CSC-like model, intratumoral injection of GD55 markedly suppressed the growth rate of xenograft tumors and induced higher levels of cell death and necrosis within the tumor tissues. Our results demonstrate that GD55 infection exerts strong anticancer effects on prostate CSC-like cells in vitro and in vivo, and has a potential to be used in the clinical therapy of PCa.