Digital polymerase chain reaction ( PCR) has been experiencing a rapid development during the past few years. Comparing with the traditional real-time quantitative PCR ( RT-qPCR) , using the same primer and probe, the accuracy for the absolute quantification of target gene is significantly improved. The development of digital PCR is directly related to the development of microfluidics. The integrated fluid circuit is an early combination of the microfluidics and digital PCR, which has a complicated fabrication process with high cost. Recently, researchers are trying to apply the droplet microfluidics in digital PCR, and the droplet microfluidic chip is able to generate millions of droplets within a short time. Each of these droplets containing no more than one target gene is a reaction chamber during the amplification process. After amplification, each droplet is tested to achieve the absolute quantification of the target gene. This paper reviews the recent progresses of droplet digital PCR, and the applications of droplet digital PCR in biological, medical and environmental fields.

wearable microfluidics have wide applications in medical, sports and military field. with the help of wearable microfluidics, through the direct physical contact between the chip and skin, the pH, glucose, lactate, sodium/potassium, calcium and heavy metal in the body fluid can be detected from sweat, tear and saliva without puncture for blood. And these information are of great importance for the monitoring of vital signs and disease diagnosis. This paper introduced the most recent development and applications of the wearable microfluidics in the body fluid testing and drug delivery. The up-to-date research development for the drug delivery using wearable microfluidics was also briefly introduced in this article. The forecast of the emerging trend for wearable microfluidics and discussion of potential technique barriers was also provided at the end of this article.

Objective To explore the diagnosis and therapy experience of vascular ring combined with tracheal compression in infants and neonates.Methods Sixteen cases(including 7 boys and 9 girls,aged 1 day to 12 months)with vascular ring combined with tracheal compression hospitalized in Guangdong General Hospital from Jun.2004 to Dec.2009 were enrolled.In these 16 children,13 cases had congenital heart malformations.All children underwent X-ray,echocardiography and spiral computed tomography examination.Nine cases received bronchoscopy study.Fifteen cases performed surgical division of vascular ring with cardiopulmonary bypass and 1 case underwent vascular ring division and tracheoplasty.Eleven cases received management of congenital heart defect simultaneously.Results Vascular ring anomalies included pulmonary artery sling in 5 children,right aortic arch-left ligmentum/aberrant left subclavian artery in 8 cases,double aortic arch in 1 case,innominate artery compression in 1 case,and pulmonary sling combined with right aortic arch-aberrant left subclavian artery in 1 case.There were 2 ring-sling complex cases in this study.The diagnosis of vascular ring were correctly made by echocardiography in 7 children and made by spiral computed tomography in all 16 cases.Two cases combined with tracheal ring died.In the follow-up study of 11 cases,5 cases were still vulnerable to wheezing.Conclusions The common presentation of tracheal compression in infants and neonates associated with vascular ring are tachypea,stridor,and dyspnea.Multi-slices spiral computed tomography is an important imaging modality.Surgical divisions of vascular ring are safe procedure in most cases and tracheal compression can be relieved by this operation.In patients with severe tracheal stenosis,tracheoplasty should be recommended.

Ductus Arteriosus ; abnormalities ; Fatal Outcome ; Female ; Heart Septal Defects, Atrial ; etiology ; Humans ; Infant, Newborn ; Rubella Syndrome, Congenital ; complications

OBJECTIVETo analyze 68 pediatric cases with functional univentricle heart who underwent bidirectional Glenn procedure during from April 1998 to December 2005.

METHODSThere were 47 males and 21 females in this group, aged from 5 months to 14 years old and weighed from 6.7 to 30.0 kg. Among them, 39 cases were received bidirectional Glenn procedure on the right side, 13 cases on the left side and 16 cases on both sides. Three cases had the pulmonary artery banded; one case had the pulmonary artery ligated;one case had the original A-P shunt cut off; six cases had the PDA ligated; four cases had the MAPCAs cut off; one case had TAPVC corrected contemporarily; two cases of PAPVC were also corrected; four cases had the atrial-ventricular valve repaired.

RESULTSThree cases died. The mortality was 4.4%. The mean post-operative pressure of super vena cava was (15.9 +/- 2.4) mm Hg (1 mm Hg = 0.133 kPa), higher than the pre-operative one (8.3 +/- 1.8) mm Hg (P < 0.01). The mean post operative SpO(2) was (89.3 +/- 4.2)%, higher than the pre-operative one (78.4 +/- 6.0)% (P < 0.01).

CONCLUSIONSBidirectional Glenn procedure is of satisfied effect on surgical treatment for functional univentricle heart. The persistent forward flow from pulmonary artery should be reserved in bidirectional Glenn procedure.

Adolescent ; Child ; Child, Preschool ; Female ; Follow-Up Studies ; Fontan Procedure ; methods ; Heart Defects, Congenital ; surgery ; Humans ; Infant ; Male ; Retrospective Studies ; Tricuspid Atresia ; surgery

Rotundine (1 micromol L(-1)) was incubated with a panel of rCYP enzymes (1A2, 2C9, 2C19, 2D6 and 3A4) in vitro. The remained parent drug in incubates was quantitatively analyzed by an Agilent LC-MS. CYP2C19, 3A4 and 2D6 were identified to be the isoenzymes involved in the metabolism of rotundine. The individual contributions of CYP2C19, 3A4 and 2D6 to the rotundine metabolism were assessed using the method of total normalized rate to be 31.46%, 60.37% and 8.17%, respectively. The metabolites of rotundine in incubates were screened with ESI-MS at selected ion mode, and were further identified using MS2 spectra and precise molecular mass obtained from an Agilent LC/Q-TOF-MSMS, as well as MS(n) spectra of LC-iTrap-MS(n). The predominant metabolic pathway of rotundine in rCYP incubates was O-demethylation. A total 5 metabolites were identified including 4 isomerides of mono demethylated rotundine and one di-demethylated metabolite. The results also showed that CYP2C19, 2D6 and 3A4 mediated O-demethylation of methoxyl groups at different positions of rotundine. Furthermore, the ESI-MS cleavage patterns of rotundine and its metabolites were explored by using LC/Q-TOF-MSMS and LC/iTrap-MS(n) techniques.
Analgesics, Non-Narcotic ; metabolism ; Aryl Hydrocarbon Hydroxylases ; metabolism ; Berberine Alkaloids ; metabolism ; Chromatography, Liquid ; Cytochrome P-450 CYP1A2 ; metabolism ; Cytochrome P-450 CYP2C19 ; Cytochrome P-450 CYP2C9 ; Cytochrome P-450 CYP2D6 ; metabolism ; Cytochrome P-450 CYP3A ; metabolism ; Cytochrome P-450 Enzyme System ; metabolism ; Dopamine Antagonists ; metabolism ; Humans ; Isoenzymes ; metabolism ; Methylation ; Recombinant Proteins ; metabolism ; Spectrometry, Mass, Electrospray Ionization

OBJECTIVETo build animal model of skull defect for the basic research of skull defects reconstruction with autogenous cranial bone dust and its relevance to the clinical application.

METHODSThree full-thickness parietal skull defects (A, B and C) were created in 30 New Zealand white rabbits. The size of all the defects was 1 cm in diameter. The defect A was left untreated as control. Defects B and C were reconstructed with autogenous cranial bone dust. Two pieces of pyroxylin membrane were placed on the top and bottom of the defect B. every 10 rabbits were killed for analysis at 4, 8, 12 weeks after operation.

RESULTSDefect A was largely repaired with connective tissue. Defect B was repaired rapidly with newly formed cancellous bone in the early period, but the following process of the growth, remodeling and maturing of the newly-formed cancellous bone was slowly. The bone ingrowth in defect C was more mature, but could not repair the defect completely, especially in the central zone. The grafted bone dust was absorbed gradually. Active angiogenesis could be observed in the newly formed bone. For the same defect, new bone had a greater amount of calcium at 12 weeks than at 4 and 8 weeks after operation (P < 0.05). And the calcium content of new bone was higher in defect C than in defect B at 8, 12 weeks after operation (P < 0.05).

CONCLUSIONSThe osteogenesis and angiogenesis are closely related to the time and location. The pyroxylin membrane can significantly promote the formation of cancellous bone in defect with autogenous bone dust graft during the early period.

Animals ; Bone Regeneration ; Bone Substitutes ; Bone Transplantation ; Female ; Male ; Materials Testing ; Osteogenesis ; Rabbits ; Skull ; Transplantation, Autologous

Adult ; Aged ; Female ; Hemangiosarcoma ; diagnostic imaging ; Humans ; Liver Neoplasms ; diagnostic imaging ; Male ; Middle Aged ; Tomography, X-Ray Computed

Objective To establish and optimize a sensitive and specific quantitative realtime polymerase chain reaction(PCR)method for detection of hepatitis B virus covalently closed circular DNA(HBV cccDNA)in liver tissue. Methods Specific primers and probes were designed to detect HBV DNA(tDNA)and cccDNA. A series of plasmids(3.44 × 100-3.44 × 109 copies/μl)containing a full double-stranded copies of HBV genome(genotype C)were used to establish the standard curve of real-time PCR. Liver samples of 33 patients with HBV related hepatocellular carcinoma(HCC), 13 Chronic hepatitis B patients(CHB)and 10 non-HBV patients were collected to verify the sensitivity and specificity of the assay. A fraction of extracted DNA was digested with a Plasmid-Safe ATP-dependent Dnase(PSAD)for HBV cccDNA detection and the remaining was used for tDNA and β-globin detection. The amount(copies/cell)of HBV cccDNA and tDNA were measured by a real-time PCR, using β-globin housekeeping gene as a quantitation standard. Results The standard curves of real-time PCR with a linear range of 3.44 × 100 to 3.44 × 109 copies/μl were established for detecting HBV cccDNA and tDNA, and both of the lowest detection limits of HBV cccDNA and tDNA were 3.44 × 100 copies/μl. The lowest quantitation levels of HBV cccDNA in liver tissues tested in 33 HBV related HCC patients and 13 CHB patients were 0.003 copies/cell and 0.031copies/cell, respectively. HBV cccDNA and tDNA in liver tissue of 10 non-HBV patient appeared to be negative. The true positive rate was increasing through the digestion of HBV DNA by PSAD, and the analytic specificity of cccDNA detection improved by 7.24 × 102 times. Liver tissues of 2 patients were retested 5 times in the PCR for detecting cccDNA and the coefficience of variations on cycle threshold (Ct)were between 0.224%-0.609%. Conclusion A highly sensitive and specific quantitative real time PCR method for the detection of HBV cccDNA in liver tissue was established and could be used for clinical and epidemiological studies.

BACKGROUNDEchocardiography is regarded as a gold standard for measuring hemodynamic values. The ultrasonic cardiac output monitor (USCOM) is a new method for measuring hemodynamics and could provide non-invasive point of care guidance. So far, there are no published USCOM reference values for neonates, nor has USCOM's accuracy been established in this population. We aimed to determine the accuracy and clinical utility of the USCOM in healthy neonates relative to published echocardiographic data, to establish normal hemodynamic parameters that it measures, and to assess the possible role of USCOM as an alternative to echocardiography as a trend monitor.

METHODSRight and left heart hemodynamics of 90 normal neonates were measured during circulatory adaptation over the first three days of life using the USCOM and automated oscillotonometry.

RESULTSHeart rate showed a significant decline from days one to three, from 126 to 120 (P < 0.001). Systolic, diastolic and mean arterial pressures all increased significantly from 66 to 71 mmHg, 33 to 38 mmHg and 44 to 49 mmHg, respectively (P < 0.001 in each case). Right ventricular cardiac index (RV-CI) showed no change with a mean of 5.07 L × min(-1) × m(-2). Left ventricular cardiac index (LV-CI) declined from 3.43 to 3.00 L × min(-1) × m(-2) (P < 0.001). RV-CI exceeded LV-CI on all three days by a mean of 61%. The systemic vascular resistance index (SVRI), based on LV-CI, increased significantly over the three days from 1083 to 1403 dyne × sec × cm(-5) × m(2) (P < 0.001).

CONCLUSIONSNormal neonatal hemodynamic values, as indicated by USCOM, were established. LV-CI measurement showed excellent agreement with published echocardiographic studies. RV-CI was constant and exceeded LV-CI for all three days of this study. It may be falsely high due to flow velocity measurement errors arising from the pulmonary branch arteries, and may represent a limitation of the USCOM method. The progressive rise of arterial pressure and SVRI despite a declining LV-CI may indicate functional closure of the ductus arteriosus, with the greatest change occurring within the first 24 hours. Evidence of closure of the foramen ovale was not observed.

Cardiac Output ; physiology ; Female ; Heart Rate ; physiology ; Hemodynamics ; physiology ; Humans ; Infant, Newborn ; Male ; Monitoring, Physiologic ; instrumentation ; methods ; Ultrasonography ; instrumentation ; methods