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Increasing gene editing efficiencies in eukaryotic cell lines by selection of appropriate CRISPR-Cas9 reagents
Melissa L. Kelley, Žaklina Strezoska, Elena Maksimova, Hidevaldo Machado, Emily M. Anderson, Maren Mayer, Annaleen Vermeulen, Shawn McClelland, Anja van Brabant Smith

Overview of various CRISPR-Cas9 reagents to provide the highest efficiency of gene editing in your experiments.

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Knockdown of p53 by Accell self-delivering siRNA causes inhibition of p53-dependent DNA damage response in IMR-32 neuroblastoma cell line and β-amyloid toxicity in rat cortical neurons
Žaklina Strezoska, Tamara Seredenina1, Devin Leake, Annaleen Vermeulen

Here we describe how application of Accell siRNA enabled the development of a high content screening assay in IMR-32 neuroblastoma cells and a whole culture cell viability assay in primary rat cortical neurons.

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An Efficient Method for the Incorporation of Molecular Probes at Multiple/Specific sites in RNA: Levulinyl Protection for 2'-ACE ® , 5'-Silyl Oligoribonucleotide Synthesis
Xiaoqin Cheng, Shawn Begay, Randy Rauen, Kelly Grimsley, Kaizhang He, Michael Delaney

A unique method that uses a levulinate ester as a protecting group to introduce conjugates or molecular probes to virtually any location in a synthetic RNA molecule is discussed. The Levulinyl protecting group is stable in RNA synthesis conditions and can be removed without affecting the other parts of the synthesized RNA. We show the capabilities of this approach with three high-complexity synthesis examples.

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Innovative technology that enables RNAi in difficult to transfect cells
Christina Yamada, Kathryn Robinson, Allison St. Amand, Zaklina Strezoska, Greg Wardle, Anastasia Khvorova, Devin Leake

Investigations at Dharmacon have led to the development of innovative siRNA molecules that can be delivered into difficult-to-transfect cells without additional lipid reagents, virus, or instruments. This technology, Accell siRNA reagents, enables gene knockdown for functional genomic studies in a wide variety of cell types. In some instances, cells can be continuously dosed with Accell siRNAs to enable target gene knockdown for extended durations.

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Innovative technology that enables RNAi in difficult to transfect cells
Christina Yamada, Kathryn Robinson, Allison St. Amand, Zaklina Strezoska, Greg Wardle, Anastasia Khvorova, Devin Leake

Investigations at Dharmacon have led to the development of innovative siRNA molecules that can be delivered into difficult-to-transfect cells without additional lipid reagents, virus, or instruments. This technology, Accell siRNA reagents, enables gene knockdown for functional genomic studies in a wide variety of cell types. In some instances, cells can be continuously dosed with Accell siRNAs to enable target gene knockdown for extended durations.

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Modelling CLL cell and T-cell Migration in a Dynamic Circulating Model of CLL
Elisabeth Walsbyl, Paul Brennan', Guy Pratte, Andrea Buggins3, Tanja N Hartmann'', Chris Fegan1 and Chris Pepper'

We have recently developed a novel circulating model of chronic lymphocytic leukaemia (CLL) that mimics the transient interactions that take place between circulating lymphocytes and vascular endothelium. Here we show that both normal and malignant lymphocytes actively underwent transendothelial migration.

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Measuring Antitumor Effect of c-Myr-Max heterodimerization inhibitor 100258-F4 on Ovarian Cancer Cells using Cellometer Imaging Cytometry
Leo L. Chan, Jiandong Wang, Xiaoli Ma, Hannah M. Jones, Fang Song, Weiyuan Zhang, Victoria L. Bae-Jump, Chunxiao Zhou

The effect from the c-Mycinhibitor 100258-F4 was clearly observed from the G0/G1 cell cycle arrest and the increase in early and late apoptotic cell. The Cellometer method can measure fluorescent cell-based assays such as cell cycle, apoptosis, protein expression, autophagy and viability. The ability to export the data to FCS Express facilitates simple data analysis and reporting to generate results for publications.

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Quantification of Natural Killer Cell-Mediated Cytotixicity using Celigo Imaging Cytometry
Leo L. Chan, Srinivas S. Somanchi, Kelsey Rosbach, Dean A. Lee

Time-course tracking of % lysis eliminates multiple controls & the effect of non-uniform cell seeding in the final cytotoxicity calculation. The # of cells used is less than the cells needed for Release assays & Flow Cytometry. Flow cytometry & Release assays require a seeding density of 100K target cells increasing the number of effector cells to the millions. The visual observation of ADCC or CDC on the images can be convincing to conclude the functionality of antibodies or complements.

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A rapid 3D tumor spheroid analysis method using the Celigo Imaging Cytometry
Leo L. Chan, Scott Cribbes, Sarah Kessel, Olivier Dery, Catherine Swingler, Dmitry Kuksin, Tim Smith, Jean Qiu, Maria Vinci, Lisa Patterson, Sue Eccles

• Celigo Imaging Cytometer is a versatile and powerful tool for 3D tumorspheroid analysis
• Assays such as measuring optimal seeding density for forming tumorspheroids and viability measurement have been demonstrated
• Advanced assays such size measurement, growth inhibition, invasion into matrigel®, migration on to collagen and HUVEC, and tissue invasion that have been performed by manual microscope observation can now be easily quantified using the automated imaging cytometry method

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The Power Decoder simulator for the evaluation of pooled shRNA screen performance
Jesse Stombaugh, Abel Licon, Žaklina Strezoska, Joshua Stahl, Sarah Bael Anderson, Michael Banos, Anja van Brabant Smith, Amanda Birmingham, Annaleen Vermeulen

Power Decoder (written in R and Python) simulates shRNA pooled screening experiments in silico to allow for the estimation of a screen’s statistical power. Populations of shRNAs were engineered in such a way that the magnitude of depletion and enrichment was known, then using the negative binomial distribution, an in silico model was developed to successfully resemble data from an actual laboratory experiment.

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Scientific News
The Mending Tissue - Cellular Instructions for Tissue Repair
NUS-led collaborative study identifies universal mechanism that explains how tissue shape regulates physiological processes such as wound healing and embryo development.
Most Complete Human Brain Model to Date is a ‘Brain Changer’
Once licensed, model likely to accelerate study of Alzheimer’s, autism, more.
Capturing Cell Growth in 3-D
Spinout’s microfluidics device better models how cancer and other cells interact in the body.
Protein That Turns Moles Into Melanoma Cancer Identified
Moles can turn into cancer, if the genetic factors recently identified by a team of researchers at the University of Pennsylvania were not present in humans.
Scientists Grow Human Serotonin Neurons in Petri Dish
The advance could facilitate the discovery of new antidepressants and drugs for illnesses involving serotonin.
Study Details Powerful Molecular Promoter of Colon Cancers
Findings show how suppression of microRNA family of molecules leads to intestinal tumors.
From Pluripotency to Totipotency
Studies results provide new elements for the understanding of pluripotency and could increase the efficiency of reprogramming somatic cells to be used for applications in regenerative medicine.
Cancer Treatment Models get Real
Researchers at Rice Univ. and Univ. of Texas MD Anderson Cancer Center have developed a way to mimic the conditions under which cancer tumors grow in bones.
Potential Treatment for Muscular Dystrophy
A new method for producing muscle cells could offer a better model for studying muscle diseases, such as muscular dystrophy, and for testing potential treatment options.
Protein Related to Long Term Traumatic Brain Injury Complications Discovered
NIH-study shows protein found at higher levels in military members who have suffered multiple TBIs.
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