Microplate Reader Grant Recipients Microplate Luminometer My lab is interested in how outside signals such as hormones and growth factors are interpreted in the cellular signaling network. We use a variety of high-throughput experimental methods to gather information about multiple signaling pathways and perform computational analysis to identify new drug targets. We use the Veritas Microplate Luminometer with Dual Injectors for many of our experimental assays due to the sensitivity of luminescent assays combined with the high-throughput design. In particular, we rely on reporter gene assays to determine the activity of transcription factors, luminescent assays for kinase activity and cAMP levels to monitor signaling activity, and a variety of luminescent assays to characterize cell behaviors such as apoptosis and proliferation. Microplate Reader Fluorometer Microplate Multimode Reader The Modulus is in place at the Shoklo Malaria Research Unit. We tested more than 70 clinical isolates (blood samples from patients with falciparum malaria) from the clinics served by SMRU. These clinics treat almost exclusively refugees and migrant workers from Burma, primarily members of the Karen hill tribe. The isolates were grown in vitro in the presence of serial dilutions of 8 different antimalarial drugs (chloroquine, quinine, piperaquine, mefloquine, artesunate, dihydroartemisinin, lumefantrine, and doxycycline) to determine susceptibility or drug resistance. Matching plates were set up for each isolate, one assayed using a pLDH DELI absorbance-based plate reader method, and the other assayed using our SYBR green fluorescence-based method. All plates were read on the Modulus, and a randomly selected subset of these were also read using transfer to mini-tubes and the TBS-380. In short, the assay and the Modulus out-performed the DELI (less failed assays, more valid results so better capture of epidemiologic data) at far lest cost and will be their standard from now on. This also validates that top-reading can work (although I still think it is not optimal) for this purpose in a challenging setting and should make the Modulus an immediate candidate for this role. Basically, I think there will be no further debate as to the method, and attention will shift back to the choice of reader method, making it very timely. Microplate Multimode Reader set up as a Luminometer I am the Director of my university’s Molecular Biology Core Facility (MBCF), a multi-user facility dedicated to advancing molecular biology-based research and education for all interested students and personnel. The MBCF accomplishes this mission by 1) providing access to, and training in, the theory and use of instrumentation not affordable by smaller laboratories; 2) providing consultation services and technical assistance to educators and researchers; 3) performing analyses on a fee-for-service basis; and 4) offering a variety of educational experiences including classroom demonstrations, student research rotations and hands-on intensive workshops. Since the MBCF is the ONLY facility of its kind within a 90 mile radius all faculty, staff, and students (undergraduate, graduate, or medical) interested in biotechnology utilize the programs offered by the MBCF either directly or indirectly (e.g., a faculty member receiving training and bringing it to their laboratory). Further, while the MBCF was clearly developed to serve the needs of the East Tennessee State University community, it routinely hosts faculty and students from nearby colleges so that they may have access to technology not available to them otherwise. Microplate Multimode Reader set up as a Luminometer
Tanja Stoyan, Ph.D. Coordinator Undergraduate Teaching Laboratory
University of California Santa Barbara Santa Barbara, California I coordinate and teach laboratory classes for undergraduates in pharmacology and immunology. Approximately 60 students graduate from these classes each year with a B.S. in Pharmacology or a B.S. in Microbiology/Cell Biology. These are classes for seniors and many of our students go on to work in the Biotech and Pharmaceutical Industries after they graduate. We are intending to expose our students to a wide variey of biochemical techniques that apply to molecular pharmacology. Recently we have developed new exercises for our classes using Flow Cytometry. My goal is to introduce our Pharmacology and Immunology students to the field of Luminometry and to develop several exercises that use this powerful technique. I will give you a brief overview about the Pharmacology program. The lab courses require pharmacology majors to spend one day each week for two quarters performing experiments to demonstrate the site and mechanism of action of a wide variety of drugs and chemical agents using different in vitro isolated tissue preparations, as well as in vivo, using anesthetized mice and rabbits. The winter quarter lab is designed to teach common biochemical techniques that are routinely used to investigate site and mechanism of action of drugs. The series of laboratory exercises include an ELISA assay to determine the concentration of protein in a sample, a radioactive drug-receptor-binding filter assay, a pharmacokinetic study to identify metabolites following different routes of administration of a therapeutic agent, isolation of liver microsomes for drug metabolism study, including identification of p450 enzymes by gel electrophoresis and immuno-blot, the amplification of G protein cDNA using PCR and a new Flow Cytometry exercise to determine the EC50 of drugs on apoptotic Jurkat cells. In addition to the experiments described above the students are required to complete a 3-dimensional molecular modeling computer exercise teaching them how to interpret structure-activity relationship of drugs and molecular binding at drug-receptor sites. The students use various software packages to analyze their data (Excel, Sigmaplot, Flow Cytometry software). Our goal is to keep the class up to date with new technologies and instrumentation. We feel that luminometry is a very important research tool. We will be using Cell Viability and Apoptosis assays, Cell titer glo, Caspase-Glo, High Through put compound screening: p450 glo, kinase-glo plus assay, pGL4, luciferase reporter vectors, Dual-glo luciferase assay system. One assay that we are interested in developing is an assay to screen for GPRC modulators which is described in Cell Notes, Issue 16, 2006 from Promega. We would introduce a luciferase reporter plasmid into various cell lines and have the students add drugs from an 80 compound library to the cells (alternatively a series of dilutions of one single drug). We would then assay for firefly and Renilla luciferase activity using the Dual-glo assay system. The results would be measured in a 96 microplate luminometer. Another application would be to assay apoptotic activity of drugs in our compound library on Jurkat cells with a luminescent assay. A high throughput screen using Flow cytometry has been difficult to do in the class format since the time for reading the 96 well plate is too long. Microplate Luminometer Personalized Diagnosis and Treatment
of Autism Spectrum Disorders
Microplate Luminometer I run a shared instrumentation facility with a focus on training rather than on service. My research goals are aligned with the users of the facility, who bring a wide variety of topics into the lab. We have people working on projects ranging from bacterial to viral to human, doing different things from the most basic science to drug screening. Our goal is to ensure that the scientists at UCSF, as well as other UC institutions, have access to advanced instrumentation for high-throughput biology, whatever the specific topic of research. We have a number of high-throughput liquid handlers, so many projects could be applied to a Veritas luminometer. We have researchers interested in established luciferase protocols, but also expect users of the facility to develop new assays with the tools we provide. My laboratory is the ideal place for a Turner Luminometer to get the maximum possible exposure to the largest number of researchers. We have several other small instruments that we have received through university grant programs; the granting companies have been very pleased with the results. As a place of interactions on the topic of new technologies for research at UCSF, my lab is unparalleled. Microplate Luminometer Microplate Luminometer |
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