RESUMO
New methods are described that expand the scope of the Successive Ring Expansion (SuRE) with respect to synthetically challenging lactams. A protocol has been developed for use with 'unreactive' lactams, enabling SuRE reactions to be performed on subsrates that fail under previously established conditions. Ring expansion is also demonstarted on 'reactive' lactams derived from iminosugars for the first time. The new SuRE methods were used to prepare a diverse array of medium-sized and macrocyclic lactams and lactones, which were evaluted in an anti-bacterial assay against E. coli BW25113WT.
RESUMO
Flow cytometry is used extensively in stem cell investigations but there is wide variation in the methods used for data analysis between laboratories. Data analysis can be challenging in stem cell biology as there is often no clear distinction between positive and negative populations. We have undertaken a critical appraisal of factors that affect the accuracy of results in stem cell applications. We used mouse embryonic stem cells and determined the expression of three common antigens in stem cell investigations, namely CD15, CD184, and c-kit. We have compared different cell preparation methods and gating strategies and also evaluated the use of isotype controls and unstained cells as controls for the identification of positive populations. The use of a "doublet discriminator" using a side scatter area signal versus side scatter height signal dot plot to identify single cells for analysis increases the accuracy of results regardless of the method used to dissociate cells. Isotype controls can be helpful in mimicking cellular nonspecific binding of the experimental antibody reaction. Isotype controls behave differently on stem cells at different stages of differentiation. Analysis of a viable single cell population with careful selection of control cell populations increases the accuracy of results.