Antibody drug conjugates (ADCs) represent a novel method to deliver a drug to diseased cell using the specific antigen-binding capability and internalization process. Antibodies that do not internalize efficiently will have sub-optimal therapeutic results.

This webinar will introduce reactive chemistries, bright and versatile fluorophores, and powerful detection platforms. You will learn how these novel technologies can advance your ADC discovery and research.

Learn how to:

• Use  pH change to detect and measure internalization
• Perform defined conjugation with the SiteClick™ Antibody Labeling system
• Quickly and easily screen candidate antibodies for specific internalization

Most cell cycle assays amount to counting cells and/or measuring DNA content and deconvolving the resulting histogram into G1, S, G2+M, or G1, S, G2, and M.  Considerable effort has been put to correlating DNA content with RNA, protein, or specific epitopes to determine or discover additional compartments.  Here, we'll focus on flow cytometric analysis of DNA and specific epitopes.  In recent efforts, using DNA peak to mark anaphase/cytokinetic populations in a 6 parameter analysis (DNA-area, DNA-peak, Light Scatter, cyclin A2, cyclin B1, and phospho-S10-histone H3), we identify 15 compartments with more than 4 additional compartments representing apoptotic cells.

Rational approaches to fixation and optimized staining protocols were worked out years ago.  We have examined many experimental variations on the basic protocols without uncovering any major improvements that result in higher quality data.  We are working currently on a “washless” staining assay, relying on a single high dilution to minimize background staining and relying on acoustic focusing to render the sample analyzable over a short period.  The results are striking. The S/N is equivalent to a fully washed sample; the minimized handling results in better cell recovery, and improved recovery results in better definition of cell cycle compartments with low cell numbers.  Additionally, the “washless” assay provides significant labor savings.  For research use only.  Not for use in diagnostic procedures.

Learning Objectives:

• overview of fixation & staining for intracellular epitopes
• underlying logic and purpose of cell cycle analysis
• in depth, analytical protocol for multiparameter analysis backbone

One of the goals of the “3Rs principle” is to replace in vivo testing methodologies with in vitro procedures on human cells, followed by computational systems biology modelling to determine toxicological risk. System toxicology, which integrates standard toxicology together with quantitative analysis of large networks of molecular and functional changes, have demonstrated the potential to help realize the 3Rs vision. Omics and high content screening methods represent fundamental pillars of a system toxicology-based approach and at Philip Morris International R&D we have developed a large battery of HCS assays to investigate how specific substances alter the phenotype of a cell grown in submerged conditions. As more physiological cell culture model, such as organotypic 3D culture are also part of our portfolio, we invested in the development of imaging-based approaches which would enable high-content investigations of 3D bronchial tissues. In this seminar, recently developed approach for phenotypic analysis of organotypic 3D Bronchial Tissues will be presented.

Cancer research diagnostic services in pathology laboratories worldwide have been negatively impacted by a lack of fresh-frozen tissue samples, the degradation of DNA and RNA in archival tissue, and tumor heterogeneity. To get accurate and reproducible gene expression and amplification results using archival material, a more robust and accurate test adapted to current clinical research workflows is needed.

Thermo Fisher Scientific, the world leader in serving science, has developed a comprehensive portfolio of Invitrogen™ immunoassays based on Luminex® xMAP® (multianalyte profiling) technology, a high-throughput solution enabling simultaneous detection and quantitation of multiple proteins. This efficient technology has proven to be invaluable for the comprehensive study of biological systems.

For the quantification of RNA in archival material, the Invitrogen™ QuantiGene™ Plex Assay is a fast and efficient multiplex solution using branched DNA (bDNA) technology on Luminex technology–based magnetic beads. Institutions, as well as professors like Godfrey Grech, have partnered with Thermo Fisher Scientific to apply this method, and have determined that it has a wide range of potential applications in the diagnosis of tumors and is adaptable to the current diagnostic research workflow.

Professor Grech will explain the main advantages of this method, including: (1) accurate, simultaneous detection of multiple targets simultaneously, enhancing accuracy and sensitivity while minimizing use of precious patient samples, (2) exclusion of subjectivity and ambiguous results originating from image-based measurements, and (3) elimination of requirements for highly specialized facilities and human resources.

This method has a wide range of possible applications, including tumor classification with diagnostic potential and measurement of biomarkers in liquid biopsies, enabling better patient management and disease monitoring. Quantitative measurement of biomarkers in archival material is also useful in oncology research with access to libraries of clinically annotated material—allowing for the validation of potential biomarkers and their correlation with clinical research outcomes using retrospective studies.  

In this webinar, we will cover the following topics:

• The benefits of multiplexing and Luminex technologies
• The use of QuantiGene Plex Assays to accurately measure RNA from archival tumor sections
• How to achieve quantitative measurement of biomarkers and correlation with clinical outcomes in retrospective studies 
• Q&A opportunities with the experts

Join us for this scientist-to-scientist presentation to share and discuss how this powerful immunoassay platform can help you meet your research goals faster.

The 7 Steps of Protein virtual event offers both broad and in-depth content, designed to give you the information and insights you need to empower your protein research every step of the way. You will receive firsthand knowledge of the latest technologies as well as tips and tricks in key protein research application areas.

Join us live on September 7, 2017 to:

• Attend valuable webinars and poster presentations— by scientists, for scientists!—to gain insights from industry experts on how to optimize your protein biology results

• Chat live with speakers and get your questions answered in real time

• Access a wide selection resources, such as technical handbooks, white papers, how-to-use product videos, and product selection guides

• Receive free Continuing Education (CE) credits for the presentations you attend

The 7 Steps of Protein virtual event presentations:

	Step 1—Protein Expression In vitro research method for screening inhibitors of protein translation (poster presentation) Approved for CE credits Speaker: Krishna Vattem, Ph.D. See Biography/Abtract
	Step 2—Protein Isolation and Purification High capacity magnetic supports for automated antibody and epitope-tagged protein purifications (poster presentation) Approved for CE credits Speaker:  Barbara Kaboord, Ph.D. See Biography/Abstract Antibody purification: Development of two new highly efficient purification resins (poster presentation) Speaker:  Aaron McBride See Biography/Abstract Efficient and convenient enrichment of multispanning membrane proteins for proteomic studies (poster presentation) Speaker: Joanna Geddes See Biography/Abstract
	Step 3—Protein Gel Electrophoresis Choosing the right protein gel for your research application (webinar) Approved for CE credits Speaker:  Alok Tomar, Ph.D. See Biography/Abstract
	Step 4—Western Blotting Classical and modernized western blotting (webinar) Approved for CE credits Speaker:  Paul Haney, Ph.D. See Biography/Abtract Chemiluminescent western blot detection: Bright and bold detection no matter how scarce your target (webinar) Speaker:  Emily Halbrader See Biography/Abstract
	Step 5—Protein Assays and Analysis Protein concentration assays – novel, simplified and rapid techniques for quantifying proteins in solution (webinar) Approved for CE credits Speaker:  Ramesh Ganapathy, Ph.D. Biography/Abstract
	Step 6—Mass Spectrometry Harnessing the power of TMT11plex sample multiplexing and improved phosphopeptide enrichment to gain new insights into signaling pathways (webinar) Approved for CE credits  Speaker: Ryan Bomgarden, Ph.D. Biography/Abstract
	Step 7—Crosslinking and Modification Strategies for successful crosslinking and bioconjugation applications (webinar) Speaker: Greg Hermanson, Chief Technology Officer and principal at Aurora Microarray Solutions, Inc. & President of Greg T. Hermanson, Inc. See Biography/Abstract

Note: All speakers are affiliated with Thermo Fisher Scientific unless otherwise indicated.

Also, don’t miss these great offers:

No matter where you are in your protein research, we offer the products and tools you'll need to take your research further.

*Discount will apply to orders received by Life Technologies no later than December 31, 2017 or until promotional supplies are depleted, whichever comes first. Customers can use the discount only once. Discount applies to the list price in effect at the time the order is received by Life Technologies. Cannot be combined with other discounts or promotions. Offer void where prohibited, licensed or restricted by federal, state, provincial, or local laws, or regulation, or agency/institutional policy. This promotion is not open to customers in Australia, New Zealand, Singapore, India, Korea, Japan, Taiwan, Brunei, Indonesia, Malaysia, Myanmar, Philippines, Thailand and Vietnam. Please contact your local sales representative for more information. Other restrictions may apply.

**No purchase necessary. This promotion is available only to qualifying attendees of 7 Steps of Protein who earned an event certification. Free t-shirt is available to eligible participants who submit a request no later than September 30, 2017 or until gift supplies are depleted, whichever comes first. The estimated value of the t-shirt is $12.00 USD. One free gift per person. Life Technologies will fulfill t-shirt size requests whenever possible, but reserves the right to substitute sizes if necessary. Health care professionals may not participate in this promotion. Health care professional (HCP): means any individual (with a clinical or non-clinical role; whether a government official, or employee or representative of a government agency or other public or private sector organization; including but not limited to, physicians, nurses, technicians, laboratory scientists, researchers, research coordinators or procurement professionals) that in the course of their professional activities may directly or indirectly purchase, lease, recommend, administer, use, supply, procure or determine the purchase or lease of, or who may prescribe medical technologies or related services. The definition of health care professionals includes individuals involved in research activity, not just those directly involved in the provision of healthcare services. Government Officials may not participate in this promotion. The term Government Official includes anyone who acts in an official capacity for or on behalf of a government entity, department, or agency. By registering, you warrant that you are not prohibited by employment, contract, or law from accepting a gift from Life Technologies. Offer void where prohibited, licensed or restricted by federal, state, provincial, or local laws or regulation or agency/institutional policy. Other restrictions may apply. If you are located in Europe, the Middle East, or Africa,  a copy of the complete Terms and Conditions is available online here. 

For Research Use Only. Not for Use in Diagnostic Procedures.

Choosing the right path to discovery can hasten your success, but the wrong path can lead to missteps that extend the journey at the expense of time, money, and frustration.

With over 40 years dedicated to cell imaging research, we offer long-proven tools and protocols to help confidently create quality cell images the first time. In fact, Invitrogen™ imaging reagents are cited more frequently in published research than any others. Leverage our experience to enable your success and avoid costly missteps.

Whether you’re new to cell imaging, or an experienced researcher wanting to confirm your knowledge, consider these five proven steps to help ensure that your cell images are publication-ready the first time. 

Early detection of protein biomarkers is critical to the study of human disease (cancer, neurobiology, inflammation/autoimmunity, etc). Basic research scientists, as well as translational investigators seek sensitive protein quantitation tools that provide low-level detection of disease relevant protein analytes. The enzyme-­linked immunosorbant assay (ELISA) first developed in the 1970s and slightly improved upon over the years, is still considered the gold standard for specific protein quantitation. However, large sample consumption, limited sensitivity, and laborious workflows associated with ELISA methods leave room for improvement and new innovation. 

Thermo Fisher Scientific, the worldwide leader in life science tools for researchers, is unveiling an affordable new platform for the next generation of high­sensitivity, ready-to-use immunoassays. Featuring serum compatible, TaqMan™ based proximity ligation assay (PLA) and Siteclick™ antibody labeling technologies, this new assay combines the analyte specificity of high affinity antibody-antigen binding with the signal detection and amplification of real-time PCR. Not only does this assay technique allow measurement of low expressing proteins that we may not have otherwise been able to detect, but it also provides an easy method to verify gene expression at a protein level. 

In this webinar, we will address the following items related to this new platform: 

• Small sample consumption – uses 2-5 µL of sample (for example, 2uL vs 150uL for triplicate wells with other methods)  
• High-sensitivity – detect low levels of proteins with greater sensitivity than other traditional methods 
• Large dynamic range – 5 logs or greater minimizes guessing associated with sample dilutions
• Fast, easy workflow – no wash steps, 2 hours from sample to answer. Step 1: Mix samples with antibodies and incubate. Step 2: Add master mix reagents and place into real time PCR instrument.
• No proprietary instrument to purchase - Runs on any real time PCR instrument – no need for specialized equipment
• Includes intuitive free cloud-enabled software – for robust data analysis and statistical groupwise comparison
• Publishing - The importance of validation for publishing purposes
• Q & A with the expert

A number of new instruments have appeared in recent times to accommodate different applications and highlight technology advances. This webinar will focus on advances in the Invitrogen™ Attune™ NxT Flow Cytometer and outline where the technology within the Attune NxT fits in current and future applications. 

I will discuss how the core acoustic focusing technology in the instrument was developed and how it has evolved. Many flow cytometers have unique capabilities and the Attune NxT is no exception. For example, the combination of traditional hydrodynamic focusing in addition to acoustic focusing allows the Attune NxT to achieve some unique tasks such as very high sample flow rates with exceptional CVs.  Everyone knows that flow cytometry offers extreme capability with single cell analysis, but we also know how difficult it is to run very small but valuable samples.  A unique feature of the Attune NxT is that you can dilute that specimen by orders of magnitude and still run the sample in a reasonable time because the sample flow rate can be drastically increased without altering the quality of the data. This feature alone can be responsible for saving critical samples that normally may be lost.  In this regard it is pertinent to understand how and why the Attune NxT fluidics is capable of operating under these conditions and I will explain the fundamentals of the how the high sample flow technology actually works. While discussing technology, I will also explain how the Attune NxT uses a special Coherent designed laser-optics module delivery system to achieve a unique flattop (tophat) focused illumination spot that plays a significant role in the low CVs achieved under high sample flow. I will also discuss my experience with several other aspects of the core technology in the Attune NxT and show how its automated plate reader option can enhance unassisted throughput giving operators back valuable time. 

For Research Use only.  Not for use in diagnostic procedures.

Learning Objectives:

• Compare hydrodynamic and acoustic focusing in flow cytometry
• List three applications where the Attune NxT provides a unique benefit
• Describe three automated plate reader options for the Attune NxT

By following relevant data privacy laws, Thermo Fisher Scientific will take great care in processing my personal data in line with its privacy policy, which can be reviewed at www.thermofisher.com/privacypolicy.

Antibodies are used in a broad range of research and diagnostic applications for the enrichment, detection, and quantitation of proteins and their modifications.  Hundreds of thousands of antibodies are commercially available against thousands of proteins and their modifications. Unfortunately, many antibodies are poorly characterized, resulting in wasted time and cost as well as potentially flawed research conclusions.  To verify the performance and specificity of Thermo Scientific antibodies, we have created a comprehensive workflow to assess antibody specificity using immunoprecipitation combined with mass spectrometry (IP-MS).  In preliminary experiments, we screened more than 500 antibodies to nearly 100 key cancer signaling proteins expressed across 12 cultured tumor cell lines.  Approximately 70% of antibodies previously validated for immunocapture could be used to capture and identify the intended target, interacting proteins, and off-targets, and ~40% of antibodies not previously validated for IP were positive by IP-MS. To demonstrate the efficacy of these antibodies, we used a set of these antibodies to simultaneously immunocapture twelve proteins in the Akt/mTOR pathway, and then quantified the proteins and their phosphorylation in four IGF-stimulated cell lines using MS-based targeted quantification.  Benchmarking of these multiplexed IP-MS assays showed moderate correlation to quantitation with more traditional Western blotting, ELISA, and Luminex assays.

Flow cytometry is a powerful technique for the analysis of multiple parameters of individual cells within heterogeneous populations. Flow cytometers are used in a broad range of applications including immunophenotyping, fluorescent protein detection, rare event analysis, live/dead cell testing, stem cell characterization, and more. As an introduction to fluorescence, this presentation covers the following topics: the principle of flow cytometry; components of a flow cytometer; basics of fluorescence and how to interpret a dye excitation/emission spectrum; how data is displayed; compensation principles and techniques; and the types of controls recommended. This webinar is useful for new and practiced cytometer users interested in learning the general concepts and principles behind flow cytometry.

Flow cytometry is a powerful technique for the analysis of multiple parameters of individual cells within heterogeneous populations. Flow cytometers are used in a broad range of applications including immunophenotyping, fluorescent protein detection, rare event analysis, live/dead cell testing, stem cell characterization, and more. As an introduction to fluorescence, this presentation covers the following topics: the principle of flow cytometry; components of a flow cytometer; basics of fluorescence and how to interpret a dye excitation/emission spectrum; how data is displayed; compensation principles and techniques; and the types of controls recommended. This webinar is useful for new and practiced cytometer users interested in learning the general concepts and principles behind flow cytometry.

Translational investigators require powerful immunoassays for the targeted detection of proteomic biomarkers with the goal of studying human health. Cytokine profiling tools are critical to understanding how acute and chronic inflammatory conditions cancer and autoimmunity. Invitrogen™ immunoassay tools from Thermo Fisher provide sensitive and specific detection of target proteins across a range of applications and platforms, including western blotting, ELISA, and Luminex™ xMAP™ multiplexed bead-based assays.  For the latter, Luminex assays produce rich data sets (30+ analyte concentrations) from a small sample volume with sensitivity and specificity.   This presentation will highlight key characteristics of the Invitrogen Luminex multiplexed immunoassay platform and how it can accelerate quantification of relevant protein markers.  

Western blotting is a dependable, trusted method for protein detection used by thousands of labs worldwide. While the process of western blotting often seems straightforward, different variables used within the workflow can sometimes create a time consuming and complex system. A critical and instrumental step in the western workflow is the transfer of proteins from gel to membrane. Over the past several years, many different methods and devices have been introduced to help improve, accelerate and reduce hands-on time of the transfer step, regardless of the gel chemistry, membrane type or protein of interest involved. In this session we will discuss the key aspects and benefits of three primary western blotting transfer technologies: traditional wet (tank) transfer, semi-dry transfer and dry transfer.

High content imaging and analysis offers a robust, high throughput method for analyzing large numbers of cells with the benefit of the spatial and temporal demarcation afforded by fluorescence microscopy. This presentation will provide an overview of high content imaging platforms, the fluorescent labels required to automate the segmentation of cells and their subcellular domains, and newly developed fluorescent assays. Emphasis will be placed on dyes that provide an indication of cell viability, mechanisms of cell death, proliferation and prelethal toxicity. Newly developed phenotypic assays—in particular those applicable to CRISPR-Cas9 mediated genome-editing—will be discussed. Novel probes and labeling approaches to report internalization of ligands and therapeutic antibodies will also be presented with reference to high content imaging.

For many years, the Laemmli system has been the standard and most popular method used to perform polyacrylamide gel electrophoresis. Laemmli type gels are useful for a broad range of protein separations. Over the past several years new technologies and innovations have provided more options to researchers, improving on the traditional Laemmli system – optimizing sharpness and band migration, improving shelf life, increasing sample load volumes, etc. New formulations designed around in-run protein degradation reduction or various protein size ranges have also helped to advance this everyday application. We will discuss some new options for protein gel electrophoresis while covering the chemical and biological advantages of these different gel technologies.

In this presentation, we will go over some of the reasons you would want to consider using fluorescence imaging and give a brief introduction to using fluorescent probes for cell structure and organelle dynamics; basic cellular function including cell cycle, endocytosis and signaling; and finally, processes involved in cell stress and death such as apoptosis, autophagy, and oxidative stress. We will also go over fluorescent labeling tips and tricks from the Molecular Probes™ School of Fluorescence Basic Imaging Module. Part of getting good results is reviewing your less-than-perfect outcomes and figuring out what to try next time. You will learn about some of the problems you may encounter, see examples of what it looks like when things go wrong, and get hints for preventing those same problems in the future.