Droplet-based optical trapping for cell separation in mock forensic samples.

Optical tweezers have a wide range of uses for mechanical manipulation of objects in the microscopic range. This includes both living and static cells in a variety of biomedical and research applications. Single-focus optical tweezers, formed by focusing a laser beam through a high numerical aperture immersion objective, create a significant force, which enables controlled transport of a variety of different cell types and morphologies in three dimensions. Optical tweezers have been previously reported to capture and separate spermatozoa from a reconstituted simulated postcoital sample. We report herein the development of a simplified, more efficient cell transfer protocol that can separate and isolate both spermatozoa as well as leukocytes, with similar efficiencies as those previously reported. The new cell transfer method was used to separate sperm cells from a reconstituted mixture of spermatozoa and vaginal epithelial cells, with complete STR profiles developed from 50 cells with little evidence of contribution from the female contributor to the mixture. This modified protocol was then used to separate 21 samples of enriched leukocytes, with trapped cells ranging from 5 to 22 cells. Complete STR profiles were developed from as few as 10 leukocytes. Thus, with minimal sample preparation and a short trapping time, this method has the potential to provide an alternative to traditional differential extraction methods for separation of sperm:nonsperm mixtures while also providing versatility for separation of cells with differing morphologies.

A Randomized, Double-Dummy, Emergency Department-Based Study of Greater Occipital Nerve Block With Bupivacaine vs Intravenous Metoclopramide for Treatment of Migraine.

BACKGROUND: Greater occipital nerve blocks (GONB) are used increasingly to treat acute migraine. OBJECTIVE: We conducted a randomized controlled trial to determine whether GONB was as effective as intravenous metoclopramide for migraine. METHODS: This was a double-dummy, double-blind, parallel-arm, non-inferiority study conducted in 2 emergency departments (EDs). Patients with migraine of moderate or severe intensity were randomized to receive bilateral GONB with each side administered 3 mL of bupivacaine 0.5% or metoclopramide 10 mg IV, the putative standard of care. The primary outcome was improvement in pain on a 0-10 scale between time 0 and 1 hour later. To reject the null hypothesis that metoclopramide would be more efficacious in relieving pain, we required that the lower limit of the 95% CI for the difference in pain improvement between those randomized to GONB vs those randomized to metoclopramide be >-1.3, a validated minimum clinically important difference. Secondary outcomes included sustained headache relief, defined as achieving and maintaining for 48 hours a headache level of mild or none without the use of additional analgesic medication, and the use of rescue medication in the ED. RESULTS: Over a 2.5-year study period, 1358 patients were screened for participation and 99 were randomized, 51 to GONB and 48 to metoclopramide. All of these patients were included in the primary analysis. Patients who received the GONB reported mean improvement of 5.0 (95% CI: 4.1, 5.8) while those who received metoclopramide reported a larger mean improvement of 6.1 (95% CI: 5.2, 6.9). The 95% CI for the between group difference of -1.1 was -2.3, 0.1. Sustained headache relief was reported by 11/51 (22%) GONB and 18/47 (38%) metoclopramide patients (95% CI for rounded difference of 17%: -1, 35%). Of the 51 GONB patients, 17 (33%) required rescue medication in the ED vs 8/48 (17%) metoclopramide patients (95% CI for rounded difference of 17%: 0, 33%). An adverse event was reported by 16/51 (31%) GONB patients and 18/48 (38%) metoclopramide patients (95% CI for (rounded) difference of 6%: -13, 25%). CONCLUSION: GONB with bupivacaine was not as efficacious as IV metoclopramide for the first-line treatment of migraine in the ED.

Optical tweezers as an effective tool for spermatozoa isolation from mixed forensic samples.

A single focus optical tweezer is formed when a laser beam is launched through a high numerical aperture immersion objective. This objective focuses the beam down to a diffraction-limited spot, which creates an optical trap where cells suspended in aqueous solutions can be held fixed. Spermatozoa, an often probative cell type in forensic investigations, can be captured inside this optical trap and dragged one by one across millimeter-length distances in order to create a cluster of cells which can be subsequently drawn up into a capillary for collection. Sperm cells are then ejected onto a sterile cover slip, counted, and transferred to a tube for DNA analysis workflow. The objective of this research was to optimize sperm cell collection for maximum DNA yield, and to determine the number of trapped sperm cells necessary to produce a full STR profile. A varying number of sperm cells from both a single-source semen sample and a mock sexual assault sample were isolated utilizing optical tweezers and processed using conventional STR analysis methods. Results demonstrated that approximately 50 trapped spermatozoa were required to obtain a consistently full DNA profile. A complete, single-source DNA profile was also achieved by isolating sperm cells via optical trapping from a mixture of sperm and vaginal epithelial cells. Based on these results, optical tweezers are a viable option for forensic applications such as separation of mixed populations of cells in forensic evidence.

Comparative analysis of posterior fusion constructs as treatments for middle and posterior column injuries: an in vitro biomechanical investigation.

BACKGROUND: Titanium pedicle screw-rod instrumentation is considered a standard treatment for spinal instability; however, the advantages of cobalt-chromium over titanium is generating interest in orthopedic practice. The aim of this study was to compare titanium versus cobalt-chromium rods in posterior fusion through in vitro biomechanical testing. METHODS: Posterior and middle column injuries were simulated at L3-L5 in six cadaveric L1-S1 human spines and different pedicle screw constructs were implanted. Specimens were subjected to flexibility tests and range of motion, intradiscal pressure and axial rotation energy loss were statistically compared among five conditions: intact, titanium rods (with and without transverse connectors) and cobalt-chromium rods (with and without transverse connectors). FINDINGS: All fusion constructs significantly (P<0.01) decreased range of motion in flexion-extension and lateral bending with respect to intact, but no significant differences (P>0.05) were observed in axial rotation among all conditions. Intradiscal pressure significantly increased (P≤0.01) after fusion, except for the cobalt-chrome conditions in extension (P≥0.06), and no significant differences (P>0.99) were found among fixation constructs. In terms of energy loss, differences became significant P≤0.05 between the cobalt-chrome with transverse connector condition with respect to the cobalt-chrome and titanium conditions. INTERPRETATION: There is not enough evidence to support that the cobalt-chrome rods performed biomechanically different than the titanium rods. The inclusion of the transverse connector only increased stability for the cobalt-chromium construct in axial rotation.