Non-disruptively Count and Quantify Fluorescent iPS Colonies During Secondary Reprogramming: 7 min per 6-well Plate, Dual-fluorescence Whole Well Imaging Cytometry
Poster Dec 04, 2014
SC Cribbes, S Brightwell, K Kaji
Current methodologies for the detection of inducible Pluripotent Stem (iPS) reprogramming are either disruptive e.g. flow cytometry (FC) or low in throughput e.g. fluorescent microscopy (FM). Using the Celigo S Imaging Cytometer and secondary iPS reprogramming we have developed a methodology that combines the advantages of both flow cytometry and fluorescent microscopy. This approach is based on the fluorescent identification of iPS colonies that express the four reprogramming factors, Oct4, Sox2, Klf4 and c-Myc, by expression of mOrange placed after the four factors following ires and the progress of reprogramming using fluorescent detection of the pluripotency reporter Nanog-GFP+ cells within these colonies. This method can be used to not only follow the reprogramming kinetics but could also be used to examine the effect of extrinsic factors, thus, providing a strong tool to investigate molecular mechanisms of reprogramming.
Nexcelom’s Celigo S imaging cytometer has been applied to provide automated, rapid assessment of iPS reprogramming . Using f-theta optics, Celigo provides high quality, whole well images using bright field and/or fluorescent illumination. Automated segmentation and analysis provides quantitative output of iPS reprogramming based on mOrange and GFP fluorescent colony detection.
Characterization of a Type 2 diabetes-associated islet-specific enhancer cluster in STARD10 by genome editing of EndoC-βH1 cellsPoster
Genome-wide association studies (GWAS) have identified more than 100 genetic loci associated with type 2 diabetes. The majority of these are located in the intergenic or intragenic regions suggesting that the implicated variants may alter chromatin conformation. This, in turn, is likely to influence the expression of nearby or more remotely located genes to alter beta cell function. At present, however, detailed molecular and functional analyses are still lacking for most of these variants. We recently analysed one of these loci and mapped five causal variants in an islet-specific enhancer cluster within the STARD10 gene locus. Here, we aimed to understand how these causal variants influence b-cell function by alteration of the chromatin structure of enhancer clusterREAD MORE
P450 Induction in Cryopreserved Hepatocytes from PXR and CAR Nuclear Receptor Knock-out RatsPoster
The nuclear receptors pregnane X receptor (PXR) and constitutive androstane receptor (CAR) are closely related transcription factors that regulate the expression of phase I (cytochrome P450s), phase II metabolizing enzymes and transporter genes in response to xenobiotics, including prescription drugs.READ MORE
Treatment Options for Chronic Parvovirus Viremia in Pediatric Heart Transplant Patients in a Tertiary Care CenterPoster
This abstract discusses three cases of pediatric heart transplant patients who suffered from parvovirus (B19) infection. Of these patients, two ( B & C) responded well to standard intravenous Ig therapy. Patient A however, did not respond to standard treatment and was begun on subcutaneous Ig, which effectively diminished his viral load. Thus, subcutaneous Ig infusions might serve as a second line treatment for transplant patients with parvovirus who do not respond well to the standard approach.READ MORE