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    • HPLC Method Development and Transfer Utilizing Fused-Core® Particle Technologies
      HPLC Method Development and Transfer Utilizing Fused-Core® Particle Technologies David S. Bell, Ph.D. Recorded: Dec 5 2012 4:00 pm UTC 31 mins
    • Superficially porous (Fused-Core, Core-shell) particle technologies have gained acceptance in general high-performance liquid chromatography (HPLC) and ultra-high pressure liquid chromatography (UHPLC) practice over the past several years due to improved efficiency relative to comparably sized fully porous particles. The Fused-Core option has also been shown to be a superior approach toward improving column efficiency as compared to smaller porous particle (sub-2 µm, UHPLC) technologies owing to the lessened backpressure penalties that are paid for the efficiency gains. Ascentis® Express HPLC columns were initially introduced that employed Fused-Core particles with an overall 2.7 µm diameter. These columns provide efficiencies equal to sub-2 µm particles at much lower backpressures as well as superior efficiencies when compared to fully porous 3 µm phases. There are still instances, however, where the backpressure generated by a 2.7 µm particle may preclude their use and there are also situations based on available equipment or regulatory guidance where larger particles are preferred. For these reasons, a new generation of 5 µm Fused-Core columns has been developed.
      In this seminar we will provide an introduction to the new line of Ascentis Express 5 µm HPLC columns and explore several scenarios practicing analytical chemists might encounter:

      • Desire to develop method on the more efficient 2.7 µm column, but need to be able to transfer to different location or lab
      1. Can I easily transfer methods from 2.7 µm to 5 µm?
      2. Can I easily transfer methods from 5 µm to 2.7 µm?

      • Wish to transfer methods based on fully porous column technologies to Fused-Core
      1. Can this be done easily?
      2. What gains should I expect? What can be done to optimize the gains?
      3. Can I transfer both 3 µm and 5 µm fully porous particle method to Fused-Core?

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    • New! Introducing Ascentis Express 5 um HPLC Columns with Fused-Core Technology
      New! Introducing Ascentis Express 5 um HPLC Columns with Fused-Core Technology R. A Henry Recorded: Sep 21 2012 3:00 pm UTC 53 mins
    • Core-type particles are competing strongly with small porous particles to improve the speed and resolution of HPLC and UHPLC experiments. The pioneering Ascentis Express column with Fused-Core® 2.7µm particles has exploded in popularity because it operates more ruggedly at much lower pressure than current sub-2µm porous particles, yet delivers the same ultra-high performance. This unique performance has been largely attributed to very narrow particle size distribution. Fused-Core® design advantages have also become popular for LC-MS because Ascentis Express columns surpass performance of columns with 3µm porous particles and operate ruggedly at higher velocities and similar pressures.

      With new, narrow-distribution 5µm Fused-Core® particles, the same design advantages can now be realized over traditional 5µm and 3µm porous particles that remain very popular for HPLC columns. An Ascentis Express 5µm column brings 3µm performance and extreme ruggedness at 5µm pressures to your laboratory. Like Ascentis Express 2.7µm particles, the 5µm particles show flatter van Deemter plots than same-size porous particles and allow separation speed to be maximized with minimal loss of resolution. Extremely high plates per pressure are observed. The core-type 5µm design should replace porous 5µm columns in routine HPLC applications with traditional instruments, and should also compete with porous 3µm columns in many LC-MS applications. Performance will be compared to 5µm and 3µm particle columns, and examples of method transfer will be shown. Ascentis Express 5µm will be available in the same phase modifications as the original Ascentis Express 2.7µm ultra-high performance column. Highly stable columns are available in various IDs and lengths up to 25cm.

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    • HPLC of Peptides:  Speed and Resolution with Fused-Core Columns
      HPLC of Peptides: Speed and Resolution with Fused-Core Columns Hillel Brandes, Ph.D. Recorded: Jan 25 2011 7:45 pm UTC 24 mins
    • HPLC columns featuring 2.7 µm Fused-Core (superficially porous) particles with 90 Å pores demonstrate very fast separations of small molecules because of high efficiency and a flat van Deemter plot. These particles rival the efficiency of sub-2 µm totally porous UHPLC particles, but show only about one-half the backpressure. Fused-Core 2.7 µm particles with wider (160 Å) pores have been optimized for the rapid separation of peptides and small proteins. The higher efficiency and lower pressure drop of Fused-Core particles allows preparation of longer columns with very large numbers of theoretical plates. This dramatically increases the peak capacity of the column system, which facilitates qualitative and quantitative HPLC and LC-MS analysis.

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    • Using Silica HPLC Columns in HILIC Mode:  A Practical Guide
      Using Silica HPLC Columns in HILIC Mode: A Practical Guide Merlin K. L. Bicking, Ph.D. Recorded: Apr 27 2011 3:00 pm UTC 44 mins
    • Silica is a popular option for use in “HILIC” mode, but developing new methods can be difficult because the retention patterns are so different from C18. This presentation will provide some practical guidelines for using silica in a HILIC mode – with buffer/acetonitrile mobile phases. Some compound classes are retained under these conditions, and some are not. We will discuss which compounds are good candidates for this system, and summarize how to get started with your method development experiments.

      You will learn:
      •What compounds can be separated on silica in HILIC mode, and what compounds cannot be separated.
      •Starting points for separations, and suggestions for how to adjust conditions for your separation problem.
      •Advantages and limitations of silica in this mode.

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    • A Practical 2D-LC System Developed for Complex Pharmaceuticals
      A Practical 2D-LC System Developed for Complex Pharmaceuticals Lianjia Ma, Ph.D.* and Anthony Alexander, Ph.D. Recorded: Mar 23 2011 3:00 pm UTC 31 mins
    • A comprehensive two-dimensional liquid chromatograph (LC × LC) was constructed from commercially available conventional HPLC equipments. This system utilizes two independently configurable 2nd dimension binary pumping systems to deliver independent flow rates, gradient profiles and mobile phase compositions to dual Fused-Core secondary columns. Very fast gradient separations (30 seconds total cycle time) were achieved at ambient temperature without excessive backpressure and without compromising optimal 1st dimension sampling rates by using superficially porous stationary phases. A practical approach to optimize the various inter-related instrumental parameters will also be presented.

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    • Ultra-Fast LC/MS/MS in Bioanalysis Using Fused-Core Columns
      Ultra-Fast LC/MS/MS in Bioanalysis Using Fused-Core Columns Ethan R. Badman, Ph.D. Recorded: Feb 16 2011 4:00 pm UTC 39 mins
    • With recent advances in HPLC columns and LC/MS hardware, it is possible to increase the throughput of bioanalytical assays without sacrificing quality. By using fused-core columns, it is possible to decrease the run time from ~4 min to 1 min or less, without the use of UHPLC hardware. High quality methods at high flow rates (1-3 mL/min) using non-ballistic gradients as short as 20 seconds were developed that provide comparable or better performance for accuracy, precision, sensitivity, and specificity than traditional slower LC methods. Data will be presented that show that these assays meet regulatory requirements for bioanalytical work. Limitations in the ultimate speed possible for these assays will also be discussed.

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    • Rapid LC/MS/MS-Based Methods For Opiates & Benzodiazepines
      Rapid LC/MS/MS-Based Methods For Opiates & Benzodiazepines Dwight R. Stoll, Ph.D., Assistant Professor - Gustavus Adolphus College Recorded: May 18 2011 3:00 pm UTC 26 mins
    • Increasing case loads and budget and staffing cuts in forensic laboratories continue to motivate the development of higher throughput methods, particularly for confirmatory analysis of regulated intoxicants. In this work, we have focused on the development of rapid LC/MS/MS methods for the determination of nine opiates including two glucuronide metabolites, and 16 benzodiazepines, including two amino- metabolites. Here we aim to analyze both the parent compounds and important polar metabolites in a single analysis. To this end we have compared the retention of the target compounds on two different reversed-phase HPLC stationary phases: a conventional C18 type phase, and a perfluorinated phenyl (PFP or F5) phase built upon the increasingly popular Fused-Core particle morphology. We see that the F5 phase not only generally exhibits higher retention than the C18 type phase, but also exhibits very different selectivity such that the nine opiates can be nearly completely resolved in under four minutes. We find that the mixture of 16 benzodiazepines cannot be completely resolved in a reasonable (i.e., less than 20 min.) time, however we have developed a separation with no more than three overlapping peaks in an analysis time of five minutes.

      Full Title
      DEVELOPMENT OF RAPID LC/MS/MS-BASED METHODS FOR CONFIRMATORY ANALYSIS OF OPIATES AND BENZODIAZEPINES

      Authors
      SPENCER BONNERUP, D. CHRISTOPHER HARMES, TOMAS LISKUTIN, JONNA BERRY, AND DWIGHT R. STOLL

      Department of Chemistry
      Gustavus Adolphus College
      800 West College Avenue
      St. Peter, MN 56082

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    • Lung Cancer and IHC
      Lung Cancer and IHC Jeff Gordon, Cell Marque- Rocklin, CA Recorded: May 12 2017 3:00 pm UTC 67 mins
    • Lung cancer is the most commonly diagnosed non-skin cancer in the United States. Each year, over 222,000 people are diagnosed with lung cancer, and over 150,000 succumb each year to the illness, making it also the deadliest cancer in the country. With constant advancement of treatment options, the importance of accurate diagnosis and detection of lung cancer becomes more and more relevant to the survival of the patient. Immunohistochemistry has served as the catalyst for these advancements in lung cancer diagnosis. This presentation covers many of the basic science, facts, and statistics of lung cancer, as well as the utility of immunohistochemical testing with markers such as TTF-1, napsin A, desmoglein-3, and p40 in the accurate diagnosis and survival rates of lung cancer.

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    • Retention Mechanisms in HILIC Chromatography:  Robust Method Development
      Retention Mechanisms in HILIC Chromatography: Robust Method Development David S. Bell, Ph.D. Recorded: Jun 28 2012 6:00 pm UTC 55 mins
    • Hydrophilic interaction liquid chromatography (HILIC), especially in conjunction with mass spectrometry (MS), has become a powerful tool for the analysis of a wide variety of challenging analytes. Applications of the technique have increased dramatically over the past decade, especially for the analysis of polar analytes where reversed-phase chromatography suffers. HILIC conditions employ a high percentage of acetonitrile which enables facilitated solvent evaporation in LC/MS sources and thus often an increase in analyte response when compared to more aqueous based systems. The increased retention of polar analytes afforded by HILIC provides improved selectivity and decreases the impact of endogenous species, often leading to improved qualitative and quantitative analyses.

      Although HILIC has proven useful, it has also been thwarted with complications including difficulties in method development and method robustness.

      In this presentation, studies investigating the underlying retention mechanisms dominant in HILIC chromatography are presented and discussed. Along with reversed-partitioning HILIC is well known to exhibit, ion-exchange and the interplay of the dominant mechanisms are unveiled and used to develop a model of overall retention and selectivity. Interactions that operate using different stationary phase chemistries and conditions are presented. The impact of analyte polarity and charge as well as the variations caused by high percentages of organic on these physiochemical parameters are highlighted. Throughout the discussion, examples of use and misuse of HILIC are employed to illustrate these important concepts to build a solid fundamental foundation for efficient and effective use of this powerful technique.

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    • New 3D-Printable Hydrogel-Based and Particle-Based Biomaterial Ink Platforms
      New 3D-Printable Hydrogel-Based and Particle-Based Biomaterial Ink Platforms Ramille Shah and Adam Jakus Upcoming: Jun 12 2017 3:00 pm UTC 75 mins
    • 3D-printing has emerged as a very promising fabrication platform for complex tissue engineering. However, a significant limitation is the availability of biomaterial inks that can be 3D-printed into highly, biologically functional materials and structures. Join us as Professor Ramille Shah and Dr. Adam Jakus of the Shah TEAM Laboratory of The Shah Tissue Engineering and Additive Manufacturing (TEAM) Laboratory discuss their work to expand the current 3D-printable material toolbox. They will review not only the an extensive variety of tailorable, functional, and clinically friendly biomaterials and corresponding medical constructs, but also additional materials, such as metals, alloys, graphene, and ceramics.

      Topics will include:
      •Two new, distinct 3D-printable material platforms, Hydrogel Bioprinting and 3D-Painting, that have been developed and extended to create hundreds of distinct 3D-printable materials.
      • The unique and advantageous properties of the new materials
      • Specific examples of new 3D-printable material systems including, new hydrogels for liver, kidney, and ovarian tissue and organ engineering and 3D-painted Hyperelastic “Bone”, 3D-Graphene

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    • Basic Principles of Solid Phase Microextraction (SPME) Method Development
      Basic Principles of Solid Phase Microextraction (SPME) Method Development Emanuela Gionfriddo, Ph.D.; Research Associate under Prof. Janusz Pawliszyn, University of Waterloo (ON, Canada) Upcoming: May 31 2017 1:30 pm UTC 75 mins
    • For the past two decades, Solid Phase Microextraction (SPME) has represented a convenient alternative to conventional sample prep procedures. SPME allows the simultaneous extraction and enrichment of analytes of interest from a given matrix in a single step while avoiding, or drastically minimizing, the use of organic solvents and time-consuming cleanup procedures.
      Like any other analytical method, the various parameters governing the SPME process need to be carefully optimized in order to achieve robustness and sensitivity. However, certain aspects of SPME method development are often overlooked by many users, leading to unsatisfactory performance of the technique.
      This webinar will shed light into several aspects of SPME method development. The presentation will include a theoretical explanation of SPME fundamentals and practical suggestions to overcome common errors and bias encountered when using SPME.
      The webinar is divided in three main sections: 1) optimization of extraction conditions 2) matrix modifications 3) optimization of desorption conditions for gas and liquid chromatography. Each section is divided in various subsections dedicated to each parameter affecting the performance of the SPME technique. The webinar attendees will be guided through comprehensive understanding of the technology and the critical parameters that influence the extraction process with practical examples from already existing methods.

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    • Amplified Detection of Proteins and their Interactions using Duolink PLA
      Amplified Detection of Proteins and their Interactions using Duolink PLA Manpreet Mutneja, Ph.D, MBA Sr. Product Manager, Molecular Platforms Life Science Research, MilliporeSigma Upcoming: Jun 15 2017 5:00 pm UTC 75 mins
    • Understanding the movements, modifications and interactions of proteins within a cell is key to unraveling the fundamental tenets of biology. However, the low-level expression of many proteins, combined with the transient nature of their interactions and movements, makes analyzing and understanding these processes quite difficult. Duolink® PLA, which is based on the principles of the proximity ligation assay (PLA), offers a solution to overcome these hurdles and to study the actions of endogenous proteins within cells and tissues. Combining the specificity of antibodies with the sensitivity afforded by rolling circle amplification, Duolink® PLA allows you to detect, visualize, and quantitate proteins and their interactions (even single events) where they happen within cells or tissue, all without overexpression or genetic manipulation. This seminar will cover the basic assay principle and advantages of the Duolink® PLA technology, and discuss recent applications and developments of the technology that make it an excellent tool to understand the fundamental mechanisms of biology, as well as disease states. Applications of Duolink® PLA include the investigation of cellular responses to varying stimuli, receptor dimerization and signalling cascades, post-translational modifications, and regulation of protein expression. New developments include use in flow cytometry and multiplexed detection.

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    • Advanced Cell Culture Technology for Generation of In Vivo-like Tissue Models
      Advanced Cell Culture Technology for Generation of In Vivo-like Tissue Models Stefan Przyborski, PhD - Professor of Cell Technology, Durham University UK Upcoming: Jun 30 2017 2:00 pm UTC 75 mins
    • The benefits of three dimensional (3D) cell culture are widely appreciated. More cell-based technologies are now becoming available that enable researchers to preserve the native 3D structure of cells in vitro. These can be broadly divided into three areas: aggregate-based methods; hydrogels and extra-cellular matrices; and inert scaffold-based technologies. Each has strengths and weaknesses and there is no one technology that satisfies all applications. Tissues in the body are mostly composed of different cell types that are often highly organized in relation to each other. Often cells are arranged in distinct layers that enable signalling and cell-to-cell interactions. Alternatively in tumours, cancer cells form aggregates and tissue masses composed of different cell types. Recreation of these types of architecture will significantly evolve 3D cell culture to a new level where real tissue-like structures can be generated in vitro.

      This webinar will review the alternative approaches available to researchers and provide an overview of their capabilities and example applications. More sophisticated models are developing as 3D cell culture technology becomes established and accepted as a means of creating more physiologically relevant cell-based assays. Methods that are relatively straightforward to use and that recreate the organized structure of real tissues will become valuable research tools for use in discovery, validation studies, and modelling disease.

      Key areas covered:
      • 2D vs 3D cell culture debate
      • Review of alternative approaches and the development of new technologies
      • Challenges facing 3D culture methods, in terms of technologies available and methods used
      • Showcase applications where 3D technology makes a difference
      • Future perspective for 3D cell culture technology and further development

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