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This protocol uses lipofectamine to deliver base editors (i.e., dCas9 and AIDx fusion protein) and sgRNA expression vectors into Duchenne Muscular Dystrophy (DMD) patient-derived human induced pluripotent stem cells (hiPSCs). This protocol details mutation of the 5' splice site of DMD exon50 with TAM (targeted AID-induced mutagenesis) followed by amplicon-based NGS library preparation for high-throughput sequencing analysis. This protocol can be generalized for base editing in other hIPSCs and for correcting aberrant splicing associated with other genetic diseases. For complete information on the generation and use of this protocol, please refer to Yuan et al. (2018).Melanocytes, derived from neural crest cells, are involved in melanin production. This protocol describes a method to generate induced melanocytes (iMelanocytes) from human induced pluripotent stem cells (iPSCs) using a suspension culture system, which considerably improves the differentiation efficiency. The most critical parts of this protocol are the selection of a reliable iPSC line with strong potential to differentiate into melanocytes and their stemness maintenance. For complete information on the use and generation of this protocol, please refer to our Cell Reports article, Liu el al. (2019).We present a protocol to rapidly test DNA binding and cleavage activity by CRISPR nucleases using cell-free transcription-translation (TXTL). Nuclease activity is assessed by adding DNA encoding a nuclease, a guide RNA, and a targeted reporter to a TXTL reaction and by measuring the fluorescence for several h. Resveratrol The reactions, performed in a few microliters, allow for parallel testing of many nucleases and guide RNAs. The protocol includes representative results for (d)Cas9 from Streptococcus pyogenes targeting a GFP reporter gene. For complete information on the generation and use of this protocol, please refer to the paper by Marshall et al. (2018).
Several frameworks have been developed to identify essential determinants for healthcare improvement. These frameworks aim to be comprehensive, leading to the creation of long lists of determinants that are not prioritised based on being experienced as most important. Furthermore, most existing frameworks do not describe the methods or actions used to identify and address the determinants, limiting their practical value. The aim of this study is to describe the development of a tool with prioritised facilitators and barriers supplemented with methods to identify and address each determinant. The tool can be used by those performing quality improvement initiatives in healthcare practice.

A mixed-methods study design was used to develop the tool. First, an online survey was used to ask healthcare professionals about the determinants they experienced as most facilitating and most hindering during the performance of their quality improvement initiative. A priority score was calculated for every named determints for performing quality improvement initiatives with suggestions for analysing and addressing these determinants. The tool is developed for those engaged in quality improvement initiatives in practice, so in this way it helps in bridging the research to practice gap of determinants frameworks. More research is needed to validate and develop the tool further.
This paper describes the development of a tool with prioritised determinants for performing quality improvement initiatives with suggestions for analysing and addressing these determinants. The tool is developed for those engaged in quality improvement initiatives in practice, so in this way it helps in bridging the research to practice gap of determinants frameworks. More research is needed to validate and develop the tool further.Polylactic acid (PLA) is a thermoplastic and biodegradable polyester, largely derived from renewable resources such as corn starch, cassava starch and sugarcane. However, PLA is only soluble in a narrow range of solvents such as tetrahydrofuran, dioxane, chlorinated solvents and heated benzene. The limited choices of solvent for PLA dissolution have imposed significant challenges in the development of specifically engineered PLA nanofibers with electrospinning techniques. Generally, the electrospun polymeric materials have been rendered with unique properties such as high porosity and complex geometry while maintaining its biodegradability and biocompatibility for emerging biomedical applications. In this study, a new anticancer drug delivery system composed of PLA nanofibers with encapsulated paclitaxel was developed by the electrospinning of the respective nanofibers on top of a spin-coated thin film with the same chemical compositions. Our unique approach is meant for promoting strong bonding between PLA-bpotentials for emerging applications as implantable drug delivery patch in post-surgical cancer eradication.This is a personal account of using hypnosis as an adjunct to specialist palliative care (SPC) treatment approaches. After a brief systematic review of the literature, one clinician's experience is outlined illustrated by short, anonymized case histories. It argues that the approach is underused in SPC. The barriers currently restricting its routine adoption in SPC are discussed including (1) a lack of SPC clinical trials, (2) a misunderstanding of hypnosis leading to stigma, and (3) its absence from clinicians' training pathways. While the evidence base for the effectiveness of hypnosis in 'supportive care', for example, managing chemotherapy-induced vomiting, is appreciable, there is a gap in SPC. There is little data to guide the use of hypnosis in the intractable symptoms of the dying, for example, breathlessness or the distress associated with missed or late diagnosis. There are many people now 'living with and beyond cancer' with chronic symptomatic illness, 'treatable but not curable'. Patients often live with symptoms over a long period, which are only partially responsive to pharmacological and other therapies. Hypnosis may help improve symptom control and quality of life. SPC trials are needed so that this useful tool for self-management of difficult symptoms can be more widely adopted.
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