Analysis of Stem Cell Differentiation using the Operetta High Content Imaging System

Human mesenchymal stem cells (hMSCs) are multipotent cells present in the bone marrow that can replicate as undifferentiated cells. They have the potential to differentiate to lineages of mesenchymal tissues such as bone, cartilage, muscle and fat. When cultured in vitro, these cells can undergo osteogenic, chondrogenic or adipogenic differentiation depending on the culture conditions [Pittenger et al., 1999]... View PDF

Quantitative Analysis of Embryoid Bodies Using the Operetta System

Embryoid bodies (EBs) are spherical three-dimensional cell aggregates derived from embryonic stem cells (ESCs) which serve as model systems for embryonic development. The formation of EBs allows the study of cellular and molecular interactions in a three-dimensional manner in the early stages of embryogenesis…  View PDF

A Comparison of HCA ImagAmp and Conventional Indirect Immunofluorescence for High Content Analysis (HCA) Applications

HCA ImagAmp™ is an amplification system designed to provide significant enhancement of target-specific fluorescence signals and to improve sensitivity in immunofluorescencebased assays. Amplification is achieved through enzyme-mediated deposition of multiple fluorophores in close proximity to a given target… View PDF

Cytotoxicity studies on live primary human hepatocytes using Operetta

The use of cultured primary human hepatocytes in predictive in vitro studies of drug metabolism and hepatotoxicity, in different stages of the drug discovery process, has increased in recent years. Primary liver cells are recognized to be the closest and most relevant in vitro model to the human liver as they express the entire hepatic contingent of drug-metabolizing enzymes and transporter proteins... View PDF

3D Cell Invasion Analysis Using Operetta

Cell invasion is of major importance in many biological processes such as embryonic development, inflammatory responses, the repair of injured tissues, and tumor formation and metastasis [Horwitz and Webb, 2003]. During metastasis, individual cancer cells leave the initial tumor mass, enter the circulatory system and move into distinct parts of the body [Lauffenburger and Horwitz, 1996]... View PDF

Signal Amplification Strategies for Rapid and Sensitive High Content Analysis

Protein kinase C alpha (PKCα) is associated with a wide variety of cellular processes including proliferation, adhesion and motility. Over expression of PKCα appears to be associated with certain forms of cancer, and it has been identified as a biomarker for poor prognosis in breast cancer [Lønne et al., 2010]. Upon activation with phorbol esters, PKCα translocates from the cytosol to the plasma membrane [Chun et al., 1996]. For this reason, it is best studied with a high content analysis method that provides quantitative data on subcellular localization....View PDF

Texture Analysis of the Plasma Membrane

Here, we present a different approach to the analysis of plasma membrane bound PKCa signal which is independent of any membrane dye or spot accumulation. Our plasma membrane texture-based approach has been shown to be very robust, and accurately quantifies PKCa located in the ridge-like structures of membrane between neighboring cells...View PDF

Image-based Quantification of the Huntingtin Protein using the Operetta

In recent studies it was suggested that a more soluble form of Htt represents
the toxic species of Htt protein [Arrasate et al., 2004] rather than the aggregate form. An
automated measurement and analysis of Htt protein in ST Hdh Q111/111 cells offers a promising basis for the development of new therapeutics against the disease.

Here, we present a high throughput High Content Analysis (HCA) assay for the quantification
of the specific, more soluble conformation of the Htt protein which is located in the perinuclear
region... View PDF

Analysis of Cell Migration

Cell migration plays a major role in a variety of physiological events, such as immune response and wound healing. In addition, cell migration can also contribute to pathological processes including metastasis [Lauffenburger et al., 1996; Horwitz et al., 2003]. Understanding the molecular components of migration is crucial for discovering new targets to develop drugs that affect migration. Identification of new therapeutics that manipulate cell migration would benefit from recent advancements in High Content Screening (HCS) technology.

Here, we present a method to analyze cell migration using the Operetta / Harmony HCS platform and the Oris(tm) Cell Migration Assay from Platypus Technologies, LLC...View PDF

Image-based Quantification of Apoptosis

Apoptosis, the genetically encoded program that leads to the self destruction of a cell, can be induced via two main pathways: the death receptor-mediated pathway, and the mitochondrial pathway [Hengartner MO, 2000]. Induction of either will finally result in the activation of caspases, a class of intracellular cytokine proteases which are considered to be the central components of the apoptotic response...View PDF

A Multiparametric Live-Cell Cytotoxicity Analysis

Cytotoxicity is a very complex process affecting multiple pathways. The ability to measure early indicators of toxicity is an essential part of drug discovery. An important approach to the detection of compound toxicity is a multiparametric analysis at the level of individual cells using High Content Analysis (HCA) applications [Abraham et al., 2008]. Such cell-based assays, which assess several cytotoxic events in a variety of relevant pathways…View PDF

Determination of Cell Cycle Phase

The cell cycle is a ubiquitous, complex process that underlines the growth and proliferation of cells, organismal development, regulation of DNA damage repair and tissue hyperplasia as a response to injury and diseases such as cancer. The characterization of pharmacological compounds that regulate cell proliferation and division is therefore particularly important for drug discovery research…View PDF