Factors that influence quality and yield of circulating-free DNA: A systematic review of the methodology literature.

BACKGROUND: Circulating-free DNA (cfDNA) is under investigation as a liquid biopsy of cancer for early detection, monitoring disease progression and therapeutic response. This systematic review of the primary cfDNA literature aims to identify and evaluate factors that influence recovery of cfDNA, and to outline evidence-based recommendations for standardization of methods. METHODS: A search of the Ovid and Cochrane databases was undertaken in May 2018 to obtain relevant literature on cfDNA isolation and quantification. Retrieved titles and abstracts were reviewed by two authors. The factors evaluated include choice of specimen type (plasma or serum); time-to-processing of whole blood; blood specimen tube; centrifugation protocol (speed, time, temperature and number of spins); and methods of cfDNA isolation and quantification. FINDINGS: Of 4,172 articles identified through the database search, 52 proceeded to full-text review and 37 met the criteria for inclusion. A quantitative analysis was not possible, due to significant heterogeneity in methodological approaches between studies. Therefore, included data was tabulated and a textual qualitative synthesis approach was taken. INTERPRETATION: This is the first systematic review of methodological factors that influence recovery and quantification of cfDNA, enabling recommendations to be made that will support standardization of methodological approaches towards development of blood-based cancer tests.

Local inflammatory response around diffusion chambers containing xenografts. Nonspecific destruction of tissues and decreased local vascularization.

Immunoisolation of xenogeneic pancreatic islets within membrane-bound devices has been proposed as an approach to cure diabetes. We examined the local response to implanted xenografts and allografts in comparison with isografts in diffusion chambers with 0.4-microm pore membranes when implanted into epididymal fat pads of rats. These membranes prevented host cell entry into the device but did not prevent passage of large molecules such as IgG and IgM. Well-differentiated allogeneic tissues (Sprague-Dawley rat embryonic lung implanted into Lewis rats) survived for 1 year when implanted in intact devices, but similar tissues were destroyed within 3 weeks when implanted within devices with holes poked in the membrane to allow host cell contact. In contrast, xenografts (CF1 mouse embryonic lung implanted into Lewis rats) were destroyed within 3 weeks even when implanted in devices with intact membranes. The death of the xenogeneic tissues was accompanied by a severe local accumulation of inflammatory cells and a decrease in local vascularization. When isogeneic tissues (Lewis rat embryonic lung implanted in Lewis rats) were mixed with xenogeneic tissues, a local inflammatory response occurred and both iso- and xenogeneic tissues were destroyed within 5 weeks. These results suggest the possibility that xenografts are killed by local accumulation of inflammatory cells, perhaps mediated by the release of antigens from the tissues within the device and presentation by an indirect pathway. The observation that the local response to xenografts is sufficient to kill isografts complicates issues of immunoprotection, suggesting that successful immunoisolation will require membranes that not only provide protection of the encapsulated tissues from the host immune system but also have properties that diminish the release of xenogeneic antigens.

Neovascularization of synthetic membranes directed by membrane microarchitecture.

Transplantation of tissues enclosed within a membrane device designed to protect the cells from immune rejection (immunoisolation) provides an opportunity to treat a variety of disease conditions. Successful implementation of immunoisolation has been hampered by the foreign-body reaction to biomaterials. We screened a variety of commercially available membranes for foreign-body reactions following implantation under the skin of rats. Histologic analysis revealed that neovascularization at the membrane-tissue interface occurred in several membranes that had pore sizes large enough to allow complete penetration by host cells (0.8-8 microns pore size). When the vascularization of the membrane-tissue interface of 5-microns-pore-size polytetrafluoroethylene (PTFE) membranes was compared to 0.02-microns-pore-size PTFE membranes, it was found that the larger pore membranes had 80-100-fold more vascular structures. The increased vascularization was observed even though the larger pore membrane was laminated to a smaller pore inner membrane to prevent cell entry into the prototype immunoisolation device. This significantly higher level of vascularization was maintained for 1 year in the subcutaneous site in rats.

The accuracy of ST segment deviation in prehospital cardiac monitoring.

Elevation or depression of the ST segment of the 12-lead electrocardiogram is an important and established marker of myocardial ischemia or injury. The ST segment deviation in normal individuals using standard cardiac monitoring equipment in the clinical setting of prehospital transport has not been studied. The hypothesis that we tested is that ST segment changes do not occur in normal subjects on standard prehospital cardiac monitors during transport. During a simulated transport, we monitored healthy volunteers of both sexes between the ages of 20 and 30 who had a normal cardiovascular history, physical examination, and 12-lead electrocardiogram. Fourteen of 34 subjects (41%) who qualified as normal exhibited ST segment deviation on the monitor. Of these 14, 11 exhibited ST segment depression, and 3 exhibited ST segment elevation. We conclude that normal individuals can exhibit ST segment deviation on standard prehospital cardiac monitoring equipment during routine transport.

Prior occupational experience and choice of alcoholism rehabilitation counselors.

Although the use of recovering alcoholics as counselors in professional settings is increasing, there is little information on the factors important in selecting counselors. This study looks at previous occupational socialization as one such factor. On the basis of a study of alcoholism rehabilitation counselors and their supervisors in Indiana it has been concluded that such socialization is an important factor. It is recommended that prospective employers be wary of applicants without some experience in "bureaucratic" types of work experience, that is, who have not had experience in work settings or with work requirements somewhat analogous to those in the professional setting.