We report here a novel membrane transfer-based DNA detection method, in which alkaline phosphatase labeled gold nanoparticle (AuNP) probes were used as a means to amplify the detection signal. In this method, the capture probe P1, complimentary to the 3' end of target DNA, was immobilized on the chip. The multi-component AuNP probes were prepared by co-coating AuNPs with the detecting probe P2, complimentary to the 5' end of target DNA, and two biotin-labeled signal probes (T10 and T40) with different lengths. In the presence of target DNA, DNA hybridization led to the attachment of AuNPs on the chip surface where specific DNA sequences were located in a "sandwich" format. Alkaline phosphatase was then introduced to the surface via biotine-streptavidin interaction. By using BCIP/NBT alkaline phosphatase color development kit, a colorimetric DNA detection was achieved through membrane transfer. The signal on the membrane was then detected by the naked eye or an ordinary optical scanner. The method provided a detection of limit of 1 pmol/L for synthesized target DNA and 0.23 pmol/L for PCR products of Mycobacterium tuberculosis 16S rDNA when the ratio of probes used was 9:1:1 (T10:T40:P2). The method described here has many desirable advantages including high sensitivity, simple operation, and no need of sophisticated equipment. The method can be potentially used for reliable biosensings.
Colorimetry ; methods ; DNA Probes ; chemistry ; genetics ; DNA, Bacterial ; genetics ; Gold ; chemistry ; Humans ; Metal Nanoparticles ; chemistry ; Mycobacterium tuberculosis ; isolation & purification ; Nucleic Acid Hybridization ; methods ; Oligonucleotide Array Sequence Analysis ; methods

BACKGROUND:Hepatocyte transplantation has attracted more and more attention as a therapeutic measure for liver failure and genetic metabolic liver diseases.OBJECTIVE:TO evaluate the efficacy and safety of human hepatocyte transplantation in treating hepatitis B related liver failure in one case by a 2-year follow-up.DESIGN:A case-report of 2-year follow-up.SETTING:No.9 Department of Infectious Diseases,Bioengineering Research Room,the 302 Hospital of Chinese PLA.PARTICI PANT:One inpatient with hepatitis B related liver failure was selected from the 302 Hospital of Chinese PLA.and she was diagnosed according the laboratory tests.The transplanted hepatocytes were originated frOm the healthy liver of a 24-year-old man,who had signed the protocol for liver donation before death.METHODS:The hepatocyte transplantation was completed in the Department of Radiology,the 302 Hospital of Chinese PLA in December 2004.Liver was isolated to obtain human primary hepatocytes, and then cryopreserved.The hepatocytes were transplanted into recipient spleen via femoral vein after resuscitation.The clinical symptoms,changes of blood biochemical indexes,and changes of spleen MRI signals were observed before and after operation.The patient was reexamined every half a year after operation, including liver function, blood coagulation function,B-mode ultrasonography,gastroscopy and MRI,and she was followed up for 2 years. MAIN OUTCOME MEASURES:Liver function,blood coagulation function, imaging indexes, immunological indexes,complication and rejection.RESULTS:①Totally(1-2)×1010 hepatocytes were harvested,and the viability of rewarmed hepatocytes was 60%,and finally 2×109 hepatocytes were transplanted.②Two months later,the clinical symptoms of the recipient were obviously ameliorated,and serum bilirubin and aspartate aminotransferase(AST)were obviously decreased,while prothrombin activity was markedly increased.20 months later,the MRI results showed that there was hepatocyte image in spleen.Two years after operation.the total bilirubin level was 20 μmol/L,direct bilirubin level was 7 μmol/L, alanine aminotransferase was 416.75 nkat/L,AST was 533.44 nkat/L,albumin was 37 g/L,prothrombin activity was 90%,which were all obviously ameliorated as compared with those before operation(474.5 μmol/L,340.3 μmol/L,400.08 nkat/L,1 200.24 nkat/L,38 g/L,25%).The patient left the hospital 2 months later and could do light-burdened job.No complications of hydroperitonia and liver function failure, etc.were observed,and no rejection occurred.Several reexaminations by B-mode ultrasonography all indicated the further aggravations of liver cirrhosis and esophageal varices.She was admitted to hospital for twice because of esophageal varices bleeding,and cured by endoscopic variceal sclerosis therapy.CONCLUSION:Hepatocyte transplantation can ameliorate liver function without rejection,but it cannot relieve portal hypertension.

objective: To investigate the status of empathy ability and perceived social support in military college students, and to explore the relationship between empathy ability and perceived social support. Methods: A total of 472 college students were selected from 5 military academies. The Interpersonal Reactivity Index-C (IRI-C) and Perceived Social Support Scale (PSSS) were used to evaluate the empathy ability and the perceived social support ability of military college students. Results: The total scores of IRI-C and PSSS were (51. 2 ± 10. 8) and (65. 8 ±11. 5), respectively. The total scores of PSSS and its three dimensions (family support, friend support and other support) were positively correlated with the total scores of IRI-C and its three dimensions (empathy concern, perspective taking and fantasy) (r = 0. 19-0. 35, Ps ＜ 0. 01). The scores of empathy concern and perspective taking could positively predict the total scores of PSSS (β = 0. 29, P ＜ 0. 001; β = 0. 15, P ＜ 0. 001). Conclusion: It suggests that the empathy ability of military college students is at the middle level. Empathy ability could predict perceived social support in military college students.

Objective To investigate the clinical diagnostic value of circulating tumor cells (CTCs)and circulating cell-free DNA (cfDNA) in peripheral blood samples in breast cancer. Methods From July 2017 to April 2018, 47 patients with BMC (7 in stage Ⅱ, 19 in stage Ⅲ and 21 in stage Ⅳ), 24 patients with benign breast diseases and 28 healthy people were selected. After collecting peripheral blood samples, serum and blood cells were separated. The size-based high-throughput microfluidic chip was used to capture CTCs. The real-time fluorescent quantitative PCR based on Alu sequence was used to detect the length of cfDNA(247 bp, 115 bp)in the serum, and the ratio of amplified products of long and short fragments was used as the index of DNA integrity. The Mann-Whitney U test or Kruskal-Wallis H test was used to compare the differences between the groups and analyze the relationship between CTCs and cfDNA and clinical parameters of breast cancer. The ROC curve was drawn and the area under the curve (AUC) was used to evaluate the feasibility of blood cell CTCs and plasma cfDNA detection as diagnostic criteria. Results The CTCs and cfDNA of 47 BMC patients were analyzed. The CTCs and cfDNA integrity index (Alu 247/115) of BMC patients were significantly higher than those of physical examination patients[(13.98± 12.36)cells / ml vs (1.14 ± 1.35) cells / ml; 0.7687 ± 0.3868 vs 0.5094 ± 0.2456], and the difference was statistically significant(the U value was 126.5,359.0;P<0.001), the area under ROC curve of CTCs was 0.885 (95％CI: 0.805-0.965), cut-off value was 7.68/ml, sensitivity was 80.4％, specificity was 96.4％. The area under ROC curve of Alu 247/115 was 0.727(95％CI: 0.608-0.847), cut-off value was 0.431, sensitivity was 71.7％, specificity was 71.4％. The AUC of CTCs and Alu 247/115 was 0.919 (95％CI 0.854-0.984), which was higher than the single test of each indicator. Conclusions CTCs and cfDNA may be the potential biological indicators for breast cancer diagnosis. The combined detection of CTCs and cfDNA maybe improve the diagnosis rate of breast cancer patients.