|Large-scale Microarray Analysis of Protein and mRNA Level Changes in HL-60 Cells|
Pavel Simara, Irena Koutna, Stanislav Stejskal, Martina Peterkova, Petr Krontorad, Zdenek Rucka
In this study we compare methods for large-scale microarray analysis of protein and RNA level changes in HL-60 cells, responding to differentiation stimuli. Using microarrays we have found, that level of several proteins was either up- or down-regulated after cell differentiation. In some cases there was significant correlation with appropriate genes.
|A Sensitive Fluorimetric Assay for Detection of ß-Secretase Activity Using a Novel FRET Peptide Substrate|
Xudong Zhu, Xing Han, Manpreet Mann, Rich Meyer, Xiaohe Tong, Anita Hong and Vera Rakhmanova
In order to facilitate high throughput screening of AD drug candidates, we have developed a new SensoLyte™ 520 b-secretase assay kit using a fluorescence resonance energy transfer (FRET) peptide, HiLyte Fluor™ 488-Glu-Val-Asn-Leu-Asp-Ala-Glu-Phe-Lys(QXL™ 520)-OH. The longer excitation and emission wavelengths of HiLyte Fluor™488 minimize the interference from autofluorescence and absorbance of test compounds.
|Glycoprotein Labeling and Detection: Novel Click Chemistry-based Applications for gel Electrophoresis, Flow Cytometry and Fluorescence Microscopy|
Brian Agnew, Nancy Ahnert, Suzanne Buck, Scott Clarke, Courtenay Hart, Kapil Kumar, and Tamara Nyberg
We demonstrate highly-selective and sensitive labeling methods for the detection of specific glycoprotein subclasses, including cell surface N- and O-linked glycoproteins and intracellular O-GlcNAc modified proteins, utilizing the copper-catalyzed cycloaddition reaction between azides and alkynes, or click chemistry.
|Development of a Lab-on-a-Chip for the Characterization of Human Cells |
Richter, L., Stepper, C., Mak, A., Brückl, H. and Ertl, P.
Cell chips are developed to continuously monitor mammalian cell population dynamics in a non-invasive manner. In the presented work we describe the design, fabrication and characterization of a lab-on-a-chip for quantitative cell analysis.
|Label-free Identification of Microorganisms using a Contact-less Dielectric Microsensor|
Ertl, P., Richter, L., Reinthaler, A., Stepper, C., Mak, A., Kast, M., Heer, R. and Brückl, H.
Microfabricated biochips are developed to continuously monitor cell population dynamics in a non-invasive manner. In the presented work we describe the novel combination of contact-less dielectric microsensors and microfluidics to promote biofilm formation for quantitative cell analysis.
|Proteomic Profiling in Defining Chemoresistant Breast Cancer|
Chuthapisith S, Layfield R, Kerr I, Hughes C and Eremin O
This study aims to identify protein profiles in breast cancer cells as predictors of chemoresistance by using two-dimensional gel electrophoresis and MALDI-TOF peptide mass fingerprinting. Our findings provide further insights into the complex mechanisms of chemoresistance, as well as representing an attractive starting point for the identification of potential protein biomarkers to predict response to chemotherapy in breast cancer in vivo.
|Simultaneous Dual Emission Detection for Fast Kinetic BRET Assays|
Dr. Marjan Orban
Bioluminescence Resonance Energy Transfer (BRET) is a system for monitoring intermolecular interactions in vivo. The BRET2™ demo kit has been used to prove the feasibility of performing a BRET assay on the POLARstar OPTIMA microplate reader. The POLARstar OPTIMA’s internal reagent injectors for 384-well plate format combined with high-end simultaneous dual emission detection offer a unique advantage for fast kinetic assays where simultaneous emission detection at two wavelengths is required.
|DNA Methylation Analysis – Reliable Cell Characterization in Regenerative Medicine|
Uli Hoffmueller, Stephen Rapko, Udo Baron, Georg Wieczorek, Alexander Hellwag, Cornelia Krüger, Stefan Kärst, Leslie Wolfe and Sven Olek
We demonstrate that DNA methylation patterns can serve as characteristic markers to distinguish different cell types. We have identified panels of methylation markers that are specific to mesenchymal stem cells or various differentiated cell types in the mesenchymal lineage. This method of cell type identification has a number of advantages over conventional markers in that it is robust, is both qualitative and quantitative.
|Solid Phase Synthesis of a Fluorescent Peptide: Comparison of Fmoc-Lys(5-FAM)-Resin and Fmoc-Lys[5-FAM(Trt)-Resin|
Wenyu Fu, Ling Sheng, Anita Hong and Xiaohe Tong
We have prepared and performed comparison studies using two kinds of resins, Fmoc-Lys(5-FAM)-Rink Amide resin (I) and Fmoc-Lys[5-FAM(trt)]-Rink Amide resin (II), the latter contains a phenolic hydroxyl group protected with a trityl group. Syntheses were carried out under the same standard conditions and the peptides obtained showed no significant difference in purity. The results of these studies showed that resin (I) is adequate for synthesis of C-terminal fluorescent labeled peptide.