OBJECTIVE Apolyclonalantibody-basedhomogeneouschemiluminescenceimmunoas-say was developed and optimized using AlphaLISA technology for the quantitative detection of microcys-tin-LR(MC-LR)inwatersamples.METHODS Thismethodwasbasedonacompetitivemodelin which an immune complex was formed from the ingegral binding of artificial MC-LR antigen-coated lumi-nescene beads,free MC-LR standards or sa mples,antibody and biotinylated second antibody.Next sensor bead were added that approached the i mmune co mplex through biotin-streptavidin interaction. With the exciting light,the energy was passed from the sensor luminescer before a special emission light could be observed.To opti mize the reaction conditions,working dilutions of polyclonal antibody and bioti-nylated second antibody were assayed while the effect of buffer syste ms and ti me of each reaction were evaluated.RESULTS Maininfluencingfactorsoftheassaywerediscussedasworkingdilutionsofpoly-clonal antibody and biotinylated goat anti rabbit IgG,assay buffer and reacting ti me.After opti mization of reaction conditions,MC-LR AlphaLISA could be finished in 40 min,with a sensitivity of 0.006 μg·L-1 and a dynamic range of 0.006 -5 μg·L-1 .The coefficient of variation was below 10% and average recovery was 1 07.7%.Moreover,the cross reactivity rates of MC-RR and MC-RY to MC-LR were 13.2%and0.91%,respectively.CONCLUSION Thismethodishighlysensitiveandspecific,time-saving and quite suitable for high throughput determination of MC-LR water samples.

Objective To evaluate the value of diagnosis of total bile acid (TBA ) ,CEA ,CA199 ,CA72‐4 in gastric carcinoma combined detection .Methods From 2013 January to 2014 April in hospital of each stage of gastric cancer in 53 patients ,80 patients in benign gastric disease group and healthy group of 120 people ,were detected the concentration of TBA ,CEA ,CA199 respectively , the serum CA72‐4 .Results Three testing groups ,detecting indexes in patients with gastric cancer group were TBA (59 .55 ± 20 . 56)μmmol/L ,CEA (17 .26 ± 11 .69)g/L ,CA199 (82 .08 ± 6 .9)U/mL ,CA72‐4 (68 .65 ± 23 .05)U/mL ,concentrations were higher than the other two groups ,with statistically significant difference between groups (P< 0 .05) .No statistical significance of CEA , CA199 ,CA72‐4 between group differences in gastric benign disease group and the healthy control group (P>0 .05) .Conclusion TBA ,CEA ,CA199 ,CA72‐4 index can be used as the detection index of clinical judgment of gastric tumor ,with a high clinical value of combined detection indexes of gastric cancer clinical treatment and prognosis .

Objective To explore the status of iodine nutrition in 8～10 years children after universal salt iodization(USI)in the iodine deficiency area.Methods Probability proportion to size method(PPS)or simple random sampling methods were used to sample children aged 8～10 years in iodine deficiency area in the vear 1995,1997,1999,2002 and 2005, respectively.Goiter were detected by palpation and B-ultrasound, iodine concentration in salt was detected by direct titration method and that in urine by the method of As3+-Ce4+catalytic spectrophotometry.Results After USI has been implemented,the median of salt and urinary iodine tended to mcreaseand the goiter rate tended to decrease year by year.In 2005,the coverage rate of iodinated salt was elevated to 97.2%,qualified iodize salt rate was 97.1%and edible qualified iodinated salt rate was 94.3%in the whole iodine deficiency areas.The median of urinary iodine Was 227.7 μg/L 89.7%(323/360)of the population had a level of the urinary iodine over 100μg/L Goiter rate of 8～10 years children Wag decreased from 22.3%(282/1267)to 4.4%(53/1200) from 1995 to 2005.Conclusion After 10-year USI,the status of iodine nutrition in ShaJldong Province has been promoted obviously and it is in a suitable iodine nutritional status.

Objective To assess the feasibility of a novel 99Tcm labelled polypeptide analogue for interleukin-11 receptor ( IL11R) imaging in a bone metastases model for prostate cancer. Methods A novel circular polypeptide analogue of IL11 ( c( CGRRAGGSC ) ) was indirectly labeled with 99Tcm and the product was named as 99Tcm-DTPA-IL11 RR. The labeling efficiency, radiochemical purity and stability of the product were measured with paper chromatography and high performance liquid chromatography (HPLC). The biodistribution of 99Tcm-DTPA-IL11RR was investigated in 28 ICR normal mice. The organ radioactivity was measured as percentage activity of injection dose per gram of tissue ( %ID/g). The models of bone metastases from prostate cancer were established at the tibias of BALB/c nude mice bearing human prostate cancer PC-3 cells. The tumor bearing ( n= 5 ) and standard closed fracture nude mice models underwent both 99Tcm-DTPA-IL11RR and 99Tcm-methylene diphosphonate (MDP) scintigraphy study. The images were acquired at 0.5, 1,2, 4, 6, 8, 24 h after intravenous injection of the tracers. The competitive inhibition imaging was perfomed in three tumor bearing mice. One-way variance analysis was used. Results The labeling efficiency was 90.7%. The radiochemical purity of 99Tcm-DTPA-IL11RR in normal saline solution was (99.57 ±0.09)%, (99.29 ±0.18)%, (98.95 ±0.78)%, (98.67 ±0.11)%, (96.53 ±0.91)%, (95.20±0.70)%, (88.38 ±0.22)% and (36.17±1.29)% at room temperature after0, 1,2, 3, 4, 6, 8 and 24 h, respectively. The tracer radiochemical purity in the blood of ICR mice remained over 90% at 37 C for 6 h. The labeling compounds were excreted mainly through kidney. The peak uptake of bone ( ( 1.910 ±0.109) %ID/g) and liver ( (0.366 ±0.030) %ID/g) was at 4 h after injection. In the tumor bearing mice, the uptake of spine marrow and large joints of extremities was mild. The highest uptake at tumor region was at 4 h and persistent at 6-8 h after injection. The tumor to non-tumor ratios (T/NT) were 1.17 ±0.17, 2.20 ±0.29, 3.20 ±0.15, 3.67 ±0.23, 13.61±0.85, 9.45 ±0.37 and 3.33 ±0.30 at 0.5,1, 2, 4, 6, 8 and 24 h, respectively (F=621.54, P＜0.05). In the standard closed fracture models,high uptake of 99Tcm-MDP was shown at the fracture site, with no increased 99Tcm-DTPA-IL11RR uptake noted. The tumor uptake was significantly depressed after a pre-injection of the unlabeled polypeptide analogue. Conclusions The synthesis of 99Tcm-DTPA-IL11RR is stable and the labeling efficiency is high. It may be a potential molecular probe in metastatic bone imaging for prostate cancer.

OBJECTIVETo develop an ELISA-based method for analyzing biologic activities of HIV-1 integrase and for high throughput screening of integrase inhibitors.

METHODSAfter expression, renaturation and purification of integrase, the bioactivity of integrase and the inhibition of luffin-a were evaluated with an in vitro assay based on biotin-avidin EILSA and chemiluminescent substrates.

RESULT(1) The specific activity of the purified integrase was 54.92 units/mg of protein. (2)IC(50) (concentration causing 50% inhibition of integrase) of luffin-a was (0.63 +/- 0.026) micromol/L.

CONCLUSIONThe non-radioactive assay can be used for analysis of bioactivities and high throughput screening of inhibitors of HIV-1 integrase.

Catalysis ; drug effects ; Dose-Response Relationship, Drug ; Enzyme Inhibitors ; pharmacology ; Enzyme-Linked Immunosorbent Assay ; methods ; HIV Integrase ; chemistry ; metabolism ; Humans ; Kinetics ; Luminescent Measurements ; Ribosome Inactivating Proteins, Type 1 ; pharmacology ; Substrate Specificity

Oroxylum indicum was a traditional Chinese medicine. In order to study the chemical constituents from the seed of O. indicum, the chemical constituents of 80% methanol extract of seeds of O. indicum were subjected to chromatography on silica gel, Sephadex LH-20, and preparative HPLC, leading to the isolation of eleven compounds. The structures were identified by various spectroscopic data including ESI-MS, 1H-NMR and 13C-NMR data as oroxin B (1), chrysin (2), baicalein (3), neglectein (4), quercetin-3-O-β-D-galactopy ranoside (5), quercetin-7-O-β-D-glucopyranoside (6), 2α,3β-dihydroxylluPeol (7), lupeol (8), rengyol (9), β-sitostero (10), and stigmasterol (11). Among them, compound 5 were firstly obtained from O. indicum.
Bignoniaceae ; chemistry ; Magnetic Resonance Spectroscopy ; Seeds ; chemistry

ObjectiveTo look into the distribution of “iodine suitable” region in iodine-deficient areas in Shandong province and to provide a scientific basis for guiding the redesignate of iodine-deficient areas and launch scientific supply of iodine.Methods One to 3 copies of water source samples were collected in 105 existing iodine-deficient counties by village.Water iodine content was detected by arsenic-cerium catalytic spectrophotometry.The areas with water iodine content below 10 μg/L was defined as iodine-deficient areas and among 10 - 150 μg/L were “iodine suitable areas” and greater than 150 μg/L were high iodine areas.Results The research was carried out in 14 cities,105 counties,and 1337 towns.We collected 65 716 water samples.Sample recovery efficiency reached 99.8％.The median of water iodine was 5.57 μg/L.In the 1337 towns surveyed,there were 82.05％(1097/1337) of the township with water iodine median ＜ 10 μg/L,17.43％(233/1337) between 10 - 150 μg/L,and 0.52％(7/1337) ＞ 150 μg/L.Conclusions In Shandong province,the water “iodine suitable” regions are distributed scattered with considerable proportion.In iodine-deficient areas,there are areas with high water iodine,and iodine-deficient regions should be redrawn.Emphasis should be put on iodine nutritional status of residents in “high iodine and iodine suitable” regions,and iodine supplementation should be carries out scientifically.

OBJECTIVETo follow the fate of murine epidermal stem cells (ESCs) seeded in a syngeneic dermal equivalent implanted in vivo.

METHODSEmbryonic stem (ES) cells were induced in vitro to differentiate into ESCs. After stained with a fluorescent dye Hoechst 33342, these ESCs were seeded into a fibroblast-collagen-gelatin sponge complex, functioning as a dermal equivalent model, and implanted subcutaneously into 129/J mice, which were syngeneic to these stem cells. The fate of these cells was observed with HE staining, immunocytochemical staining or Van Gieson's staining.

RESULTSThese ESCs were clearly visible in the implant by fluorescent microscopy 3 weeks or longer after implantation. These cells remained viable, differentiated into hair follicle-like structure, glandular structure, and gave rise to additional structures displaying features resembling native dermis. A number of markers were expressed in the differentiated structures, including CD29 (integrin beta1 subunit) and cytokeratin 18 (CK18). No apparent rejection or severe side effects were observed at least during the 10 weeks following implantation.

CONCLUSIONNow that ESCs could survive in vivo in this dermal equivalent model, and differentiate into hair follicle-like structures as well as glandular structures, it is feasible to use these cells as seed cells in the study to fabricate dermal equivalent having the potential to develop dermal appendages.

Animals ; Cell Culture Techniques ; Cells, Cultured ; Dermis ; cytology ; transplantation ; Embryonic Stem Cells ; cytology ; Epithelial Cells ; cytology ; Mice ; Mice, Inbred Strains ; Skin Transplantation ; Stem Cell Transplantation ; Stem Cells ; cytology ; Tissue Engineering

Objective To learn the iodine nutritional status of the vulnerable population with different iodine level under the current level of iodized salt in Shandong province and to offer prevention and cure measures.Methods Five groups of vulnerable population including school children aged 8 - 10, pregnant, lactation women, infants and women of childbearing age from mountain areas ( Daiyue, Mengyin counties ) , plain ( Luxian,Gaomi counties ) and coastal (Zhaoyuan county ) of five different iodine deficient areas were investigated in 2007.The thyroids of children aged 8 - 10 were checked by palpation and B ultrasound, their edible salt iodine level was detected by direct titration. The lever of urinary iodine of vulnerable population was examined by arsenic and cerium speetrophotometry. Results The goiter rates of 8 - 10 year-old were 1.8%(9/514) and 1.2%(6/514), respectively by palpation and B-ultrasonic. The mean iodine of 501 edible salt samples was 30.95 mg/kg. The coverage rate of iodized salt was 94.6% (474/501). The rate of qualified iodized salt was 90.4% (453/501). The median of urinary iodine was 216.7 μg,/L. The urinary iodine of school children aged 8 - 10, pregnant, lactation women, infants and women of childbearing age were 234.0, 165.5, 162.4, 257.5, 233.0 μg/L, respectively. Conclusions Current iodine nutritional level is basically appropriate in all groups of vulnerable people. The current iodine content of iodized salt could meet the needs of population from different iodine deficient areas of Shandong province.

The cDNA sequence encoding pokeweed antiviral protein-II was cloned from the fresh summer leaves of phytolacca amercana by RT-PCR. The recombinant PAP-II was subcloned into the expression vector pET-28a(+) and expressed in E. coli BL21 after IPTG induction. SDS-PAGE analysis showed that the expressed PAP-II existed in the form of inclusion bodies. The purified fusion protein was obtained after a series of steps including cell break, inclusion body solubilization, protein refolding and purification through BBST NTA resin column. The non-radioactive ELISA-based HIV-1 integrase assay showed that the recombinant pokeweed antiviral protein-II and RTA were able to inhibit HIV-1 integrase to some extent (IC50 = 303 microg/mL, 220 microg/mL respectively). MTT assay showed that cytotoxicity of pokeweed antiviral protein II for HEP-G2 cells and Hela cells was in a dose-dependent manner with IC50 s of 93 microg/mL and 102 microg/mL, respectively. The results suggested that pokeweed antiviral protein-II is a potent anti-tumor candidate. The finding of integrase inhibitory activity and the discovery of cytotoxicity provide more insights into the anti-HIV and the anti-tumor activities of PAP-II.
Cloning, Molecular ; HIV Integrase ; drug effects ; HeLa Cells ; Humans ; Phytolacca americana ; genetics ; Plant Leaves ; genetics ; Plasmids ; Recombinant Proteins ; biosynthesis ; isolation & purification ; pharmacology ; Ribosome Inactivating Proteins, Type 1 ; genetics ; isolation & purification ; pharmacology