Application of Duplex Droplet Digital PCR Detection of miR-888 and miR-891a in Semen Identification.

OBJECTIVES: To establish a system for simultaneous detection of miR-888 and miR-891a by droplet digital PCR (ddPCR), and to evaluate its application value in semen identification. METHODS: The hydrolysis probes with different fluorescence modified reporter groups were designed to realize the detection of miR-888 and miR-891a by duplex ddPCR. A total of 75 samples of 5 body fluids (including peripheral blood, menstrual blood, semen, saliva and vaginal secretion) were detected. The difference analysis was conducted by Mann-Whitney U test. The semen differentiation ability of miR-888 and miR-891a was evaluated by ROC curve analysis and the optimal cut-off value was obtained. RESULTS: There was no significant difference between the dual-plex assay and the single assay in this system. The detection sensitivity was up to 0.1 ng total RNA, and the intra- and inter-batch coefficients of variation were less than 15%. The expression levels of miR-888 and miR-891a detected by duplex ddPCR in semen were both higher than those in other body fluids. ROC curve analysis showed that the AUC of miR-888 was 0.976, the optimal cut-off value was 2.250 copies/µL, and the discrimination accuracy was 97.33%; the AUC of miR-891a was 1.000, the optimal cut-off value was 1.100 copies/µL, and the discrimination accuracy was 100%. CONCLUSIONS: In this study, a method for detection of miR-888 and miR-891a by duplex ddPCR was successfully established. The system has good stability and repeatability and can be used for semen identification. Both miR-888 and miR-891a have high ability to identify semen, and the discrimination accuracy of miR-891a is higher.

[Oral controlled-release oxycodone for uterine cramping pain after cesarean section].

OBJECTIVE: To evaluate whether combined oral oxycodone hydrochloride controlled-release tablets plus paracetamol and tramadol hydrochloride tablets is better than epidural analgesia with respect to uterine cramping pain control and side effects after cesarean section. METHODS: Sixty consecutive patients scheduled for cesarean section from April to May, 2010 were randomized to either patient-controlled epidural analgesia with 0.1% ropivacaine, 0.1 µg/ml sufentanil (for postoperative 48 h) plus injected pethidine on demand (EDA group) or controlled-release oxycodone (2 × 15 mg for 1st postoperative 24 h; 2 × 10 mg for 2nd postoperative 24 h), paracetamol & tramadol hydrochloride tablets (8 × 1 tablet for postoperative 48 h) orally plus pethidine injection on demand (OXY group). Two groups were compared with respects to uterine cramping pain control when the oxytocin infusion (20 U plus 500 ml 5% glucose solution, iv. gtt within 2 h) once per day for postoperative 3 days as determined by the means of a visual analogue scale (VAS), pethidine consumption, side effects and costs. RESULTS: The EDA group experienced significant more pain than the OXY group when the oxytocin infusion was administered (mm) [50.0 (15.0, 72.5) vs 25.0 (0, 40.0), 60.0 (47.5, 72.5) vs 20.0 (0, 30.0), 35.0 (20.0, 50.0) vs 0 (0, 20.0)]. all P < 0.05). Pethidine was used for pain control in 2 patients (150 mg total) of EDA group during the oxytocin infusion whereas none of the OXY group received an injection of pethidine. There was a higher level of maternal satisfaction with a lower analgesic dose in the EDA group (80.9 ± 9.3 vs 90.0 ± 9.8, P < 0.01). The median duration of hospital stay was around 5 days in both groups. CONCLUSION: Postoperative pain control after cesarean section with the above combined regimen is superior to EDA in terms of a lower cost and a higher level of maternal satisfaction.