Development of high-performance magnetic chemiluminescence enzyme immunoassay for alpha-fetoprotein (AFP) in human serum.

BACKGROUND: A high-performance chemiluminescence enzyme immunoassay (CLEIA) for alpha-fetoprotein (AFP), a tumor marker for the diagnosis of hepatocellular carcinoma (HCC), was constructed by using magnetic particles (MPs) as both the immobilization matrix and separation tools. METHODS: A double sandwiched immunocomplex was formed through the reaction among anti-fluorescein isothiocyanate (FITC) antibody coated MPs, FITC-labeled anti-AFP antibody, AFP antigen, and alkaline phosphatase (ALP)-labeled anti-AFP antibody. The subsequent chemiluminescence reaction of ALP with 4-methoxy-4-(3-phosphate-phenyl)-spiro-(1,2-dioxetane-3,2'-adamantane) (AMPPD) gave light intensity that was directly proportional to the amount of analyte present in the samples. The effects of several physicochemical parameters, including the concentration of FITC-labeled anti-AFP antibody, the dilution ratio of ALP-labeled anti-AFP antibody, the volume of MPs and substrate, the immunoreaction time and other relevant variables upon the immunoassay were studied and optimized. RIA and microplate CLEIA were used as comparison methods. RESULTS: The proposed method had a sensitivity of 3.0 ng/ml, low cross reactivities, and an assay time of 1 h. The linear range was 0-1200 ng/ml through using MPs and is useful for samples with extremely high AFP concentrations without dilution while avoiding the hook effect. The intra- and inter-assay precision was <3% and <5%. The present method has been successfully applied to the detection of AFP human serum with recoveries from 90 to 108%, and showed a good correlation with the commercially available AFP RIA kit. CONCLUSIONS: This proposed assay provided apparent advantages over microplate CLEIA and RIA, and facilitated the development of high-throughput screening and automated operation systems in the clinical practice.

Clinical application of 64-slice spiral CT in the diagnosis of the Tetralogy of Fallot.

OBJECTIVE: To explore the clinical application of 64-slice spiral CT in the diagnosis of the Tetralogy of Fallot. METHODS: Eighteen patients with diagnosed Tetralogy of Fallot underwent cardiac CT angiography with 64-slice CT (CTA). Two- and three-dimensional images were used for diagnosing in all cases by means of MPR (coronal, sagital and oblique), cMPR, MIP and VRT. RESULTS: All patients had ventricular septal defect, pulmonary stenosis and hypertrophy of right ventricle. The morphologic features of Tetralogy of Fallot were equal to echocardiography. Patent ductus arteriosus (PDA) were detected on eight patients. Main coronary artery branches and partial sub-branches were visualized in all patients. Abnormal coronary arteries were found in seven cases: both left and right main coronary artery arising from the right aortic sinus one case, right main coronary artery and circumflex (CX) arising from the right aortic sinus one case, left anterior descending (LAD) and CX arising from left aortic sinus directly one case, left and right coronary artery arising from back and front of the aortic sinus, respectively, three cases. Pulmonary artery branch stenosis was found in 12 cases and branch pulmonary artery dilation noticed in 1 case. Double superior vena cava was also found in one case. According to the surgery the diagnostic accuracy of CT and Echocardiography was 95.45 and 83.33%, respectively. CONCLUSION: Two- and three-dimensional 64-slice CTA not only show the overall anatomical structure of the heart, but also show coronary and pulmonary arteries. With these results, evaluation of coronary anomalies and pulmonary artery stenosis with 64-MSCT is extremely valuable for planning the operative procedure on the patients with Tetralogy of Fallot.

[The application of multi-slice CT dynamic enhancement scan in the diagnosis and treatment of colonic lymphomas].

OBJECTIVE: To discuss the value of multi-slice CT dynamic enhancement scan in the diagnosis and treatment of colonic lymphomas. METHODS: 16 patients with colonic lymphomas underwent multi-slice CT dynamic enhancement scans, images of axial and reconstructive images of VR, MPR and CTVE were analyzed, patients were respectively diagnosed. RESULTS: Appearances of primary colorectal lymphomas were categorized into focal and diffuse lesions. Focal and diffuse lesions were 6 and 10 patients, respectively. The accuracy rate of diagnosis was 87.5%. CONCLUSION: MSCT dynamic scan has distinctive superiority in diagnosis and treatment of colonic lymphomas.

Chinese expert consensus workshop report: Guidelines for thermal ablation of primary and metastatic lung tumors.

Although surgical resection is the primary means of curing both primary and metastatic lung cancers, about 80% of lung cancers cannot be removed by surgery. As most patients with unresectable lung cancer receive only limited benefits from traditional radiotherapy and chemotherapy, many new local treatment methods have emerged, including local ablation therapy. The Minimally Invasive and Comprehensive Treatment of Lung Cancer Branch, Professional Committee of Minimally Invasive Treatment of Cancer of the Chinese Anti-Cancer Association has organized multidisciplinary experts to develop guidelines for this treatment modality. These guidelines aim at standardizing thermal ablation procedures and criteria for selecting treatment candidates and assessing outcomes; and for preventing and managing post-ablation complications.

Development of magnetic particle-based chemiluminescence enzyme immunoassay for the detection of 17beta-estradiol in environmental water.

In the present work, a simple, fast, and highly sensitive chemiluminescence enzyme immunoassay for 17beta-estradiol (E2) in environmental water samples was developed, using magnetic particles (MPs) labeled with secondary antibody as both the immobilization matrix and the separation tools. The specific anti-E2 polyclonal antibody (PcAb) was produced against a conjugate of estradiol-bovine serum albumin. The specificity of the anti-E2 antibody was studied. The results showed that the antibody did not cross-react with the structurally related endocrine-disrupting compounds, including estrone, ethinyl E2, estriol, E2-17-glucuronide, E2-3-sulfate-17-glucuronide, androstenedione, and dihydrotestosterone. The water samples were pretreated with solid-phase extraction using C18 cartridges for the removal of matrix effects. Several physicochemical parameters including the dilution ratios of E2-6-horseradish peroxidase conjugate and anti-E2 PcAb, immunoreaction time, volume of chemiluminescent substrate and MPs, chemiluminescence reaction time, and pH of assay solution were studied and optimized. At optimal experimental conditions, it was found that the proposed method exhibited high performance with detection limit of 2.0 pg/mL, linear range of 20-1,200 pg/mL, and total assay time of 45 min. Both inter- and intra-assay coefficient of variation were less than 10%. The average recoveries of three different spiked concentration samples ranged from 86.3% to 108%. The method was successfully applied to the determination of E2 in river, waste, and tap water, and showed a good correlation with the commercially available radioimmunoassay kit.

Magnetic particle-based chemiluminescence enzyme immunoassay for free thyroxine in human serum.

A magnetic particles-based chemiluminescence enzyme immunoassay with high sensitivity, specificity, rapidity, and reproducibility was developed for the determination of free thyroxine in human serum. A competitive assay has been proposed with horseradish peroxidase labeled thyroxine analog. The immunomagnetic particles coated with anti-fluorescein isothiocyanate antibody was used as dispersed solid phase and separation means for the immunoassay. Experimental conditions, such as temperature, the volume of magnetic particles and substrate, incubation time, dilution ratio and other relevant variables upon the immunoassay have been examined and optimized. The proposed method exhibited high performance which the linear range was 1.59-122 pmol L(-1) and the detection limit was 0.25 pmol L(-1). A coefficient of variance of less than 15% was obtained for both intra-assay and inter-assay precision. The present method has been successfully applied to the analysis of free thyroxine in human serum. The diagnostic accordance rate of the method for normal serum, hyperthyroidism and hypothyroidism are satisfactory. Good correlations were obtained between the results by the proposed method and the commercial radioimmunoassay kit. The present method exhibits good potential in the fabrication of FT4 diagnostic kits which could be used in the clinical analysis and facilitated the development of automated operation systems in the clinical practice.

Development of a sensitive, rapid, biotin-streptavidin based chemiluminescent enzyme immunoassay for human thyroid stimulating hormone.

A highly sensitive "two-site" chemiluminescent immunoassay specific for human thyroid stimulating hormone (TSH) was developed. The signal amplification was achieved via a biotin-streptavidin system (BSAS). The HRP-luminol-H(2)O(2) chemiluminescent system with high sensitivity was chosen as the detection system. Biotinylated anti-TSH monoclonal antibody (MAb) and HRP-labeled streptavidin were first synthesized. Then the signal amplification was achieved through the interaction between the biotinylated anti-TSH MAb and the HRP-streptavidin conjugate. The light intensity developed was in proportion to the TSH present in the samples. The assay showed little cross-reactivity with three other glycoprotein hormones (human chorionic gonadotropin (HCG), luteinizing hormone (LH), follicle stimulating hormone (FSH)) due to the high specificity of the antibody. The working range for human thyroid stimulating hormone was 0.1-40 mU L(-1). Both the intra-assay and inter-assay coefficients of variation were less than 10% for the BSAS based chemiluminescent enzyme immunoassay (CLEIA). The proposed assay had a sensitivity of 0.01 mU L(-1) which was 10-fold higher than the HRP-MAb conjugate based TSH immunoassay. Thus the higher sensitivity facilitated the clinical testing for thyroid states. The effects of several reaction parameters, such as incubation time, temperature, and reaction volume of the method, were also studied. This method has been successfully applied to the evaluation of TSH in human serum. Compared with the commercial enzyme chemiluminescent immunoassay, the correlation was satisfied.