<h3>Two Dimensions, One Solution: Advancing Oligonucleotide Analysis with 2D-LC</h3><h4>Why Consider 2D-LC for Oligonucleotide Analysis?</h4>In oligonucleotide analytics, achieving reliable and precise results is not optional—it is fundamental. These biomolecules are inherently complex, often containing truncated sequences, structural variants, and a wide range of impurities. Such complexity frequently exceeds the separation capabilities of conventional one-dimensional liquid chromatography (1D-LC). As a result, critical components may coelute or remain unresolved, limiting analytical confidence.Two-dimensional liquid chromatography (2D-LC) offers a powerful alternative by significantly enhancing separation performance. By combining two complementary separation mechanisms, 2D-LC improves resolution, sensitivity, and overall analytical reliability. This enables a much deeper understanding of oligonucleotide samples and their compositional intricacies.<blockquote><h4>Previous parts from the Oligos Made Easy series</h4><ul><li>Part 1: <a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/article/6808">Overview</a></li><li>Part 2: <a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/article/6846">AEX</a></li><li>Part 3: <a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/article/6880">IP-RP</a></li><li>Part 4: <a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/article/6912">HILIC</a></li><li>Part 5: <a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/article/6944">SEC</a></li><li>Part 6: <a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/article/6982">Column Navigator</a></li><li>Part 7: <a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/article/7012">Large Scale AEX</a></li><li>Part 8: <a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/article/7045">Sepapure oliGO</a></li></ul></blockquote><h4>Moving Beyond the Limits of 1D-LC</h4>For many years, 1D-LC has served as a standard tool for biopharmaceutical characterization. However, its inherent limitations become apparent when dealing with highly complex mixtures. The selectivity of a single chromatographic dimension is restricted, and the lack of orthogonality in retention mechanisms often results in insufficient separation. As sample complexity increases, the resolving power of 1D-LC becomes inadequate.<img src="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/KNAUER_Figure_1_2_D_LC_Principle_Graphic_by_KNAUER_43e04d32a2.jpg" alt="KNAUER: Figure 1 - 2D-LC Principle (Graphic by KNAUER)." srcset="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/thumbnail_KNAUER_Figure_1_2_D_LC_Principle_Graphic_by_KNAUER_43e04d32a2.jpg 245w," sizes="100vw" width="1291" height="391">Two-dimensional LC addresses these limitations by coupling two chromatographic separations, which can be either similar or fundamentally different. Each dimension targets different physicochemical properties of analytes, enabling the resolution of compounds that would otherwise overlap. In essence, 2D-LC introduces an additional level of separation capacity, allowing analysts to disentangle coeluting species that remain unresolved in a single dimension.An important practical consideration is that the overall analysis time is governed by the second dimension. Because fractions from the first dimension must be processed sequentially, the second dimension must operate rapidly and efficiently. High-speed separations in the second dimension are therefore essential to maintain system performance and data quality.<h4>Architecture of a 2D-LC System</h4>At its core, a 2D-LC system relies on precise coordination between its components. The setup typically consists of two chromatographic columns, two pumping systems, and a switching valve that enables fraction transfer between dimensions. The sample is introduced through an autosampler and first separated on the primary column. Fractions of interest are then transferred into a loop via a switching valve and subsequently introduced into the second column for further separation.<img src="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/KNAUER_Figure_2_2_D_LC_System_Configuration_Example_Graphic_by_KNAUER_f5747adf3a.jpg" alt="KNAUER: Figure 2 - 2D LC System Configuration Example (Graphic by KNAUER)." srcset="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/thumbnail_KNAUER_Figure_2_2_D_LC_System_Configuration_Example_Graphic_by_KNAUER_f5747adf3a.jpg 245w," sizes="100vw" width="859" height="517">The valve plays a crucial role in this process, acting as the interface between both chromatographic dimensions. Its function is to alternately collect fractions from the first dimension and inject them into the second dimension under high-pressure conditions. Various valve configurations can be employed, including ten-port or eight-port valves with dual-loop arrangements, or synchronized six-port valves. Regardless of configuration, the underlying principle remains the same: while one loop is being filled, the other is being flushed into the second dimension.<img src="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/KNAUER_Figure_3_Schematic_figure_of_ten_port_left_and_eight_port_valve_right_2_position_setup_for_2_D_LC_Graphic_by_KNAUER_9209661ea8.jpg" alt="KNAUER: Figure 3 - Schematic figure of ten-port (left) and eight-port valve (right) 2-position setup for 2D-LC (Graphic by KNAUER)." srcset="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/thumbnail_KNAUER_Figure_3_Schematic_figure_of_ten_port_left_and_eight_port_valve_right_2_position_setup_for_2_D_LC_Graphic_by_KNAUER_9209661ea8.jpg 245w," sizes="100vw" width="1287" height="406">This alternating process requires precise timing. The second-dimension separation must be completed before the next fraction is ready for transfer. Consequently, synchronization between flow rates, loop volumes, and switching intervals is essential for maintaining chromatographic integrity.<img src="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/KNAUER_Figure_4_Schematic_figure_KNAUER_eight_port_2_position_valve_for_2_D_LC_Graphic_by_KNAUER_f9a5f06f5f.jpg" alt="KNAUER: Figure 4 - Schematic figure KNAUER eight-port 2 position valve for 2D-LC (Graphic by KNAUER)." srcset="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/thumbnail_KNAUER_Figure_4_Schematic_figure_KNAUER_eight_port_2_position_valve_for_2_D_LC_Graphic_by_KNAUER_f9a5f06f5f.jpg 245w," sizes="100vw" width="756" height="391"><h4>Applying 2D-LC to Oligonucleotide Analysis</h4>In oligonucleotide workflows, 2D-LC combines two complementary separation strategies to address sample complexity. The first dimension is typically used for bulk separation based on properties such as size, charge, or polarity. Techniques such as size-exclusion chromatography (SEC), anion-exchange chromatography (AEX), ion-pair reversed-phase chromatography (IP-RP), hydrophilic interaction chromatography (HILIC), or mixed-mode chromatography (MMC) are commonly applied. This step reduces sample complexity and can also serve as an online desalting stage, removing salts and buffer components that could interfere with downstream detection, particularly when mass spectrometry is used.Following this initial separation, selected fractions are transferred to the second dimension. This transfer can be performed either selectively or comprehensively, depending on the analytical goal. The second dimension is typically optimized for high-resolution separation, allowing detailed characterization of closely related species such as truncated oligonucleotides, sequence variants, or chemically modified impurities. When coupled with mass spectrometry, this setup enables accurate identification and sequencing.A key advantage of 2D-LC lies in the flexibility of combining different stationary phases and mobile phase conditions.&nbsp;Numerous separation combinations are possible, including:<ul><li>SEC×AEX</li><li>HILIC×IP-RP</li><li>IEC×IP-RP</li><li>IP-RP×IP-RP</li><li>AEX×HIC</li><li>MMC×SEC,</li><li>and even chiral×RP systems.&nbsp;</li></ul>This adaptability allows tailored workflows for specific analytical challenges.<h4>Heart-Cutting vs. Comprehensive 2D-LC</h4>When implementing 2D-LC, two primary transfer strategies are commonly used: heart-cutting and comprehensive transfer. Each approach offers distinct advantages depending on the analytical objective.Heart-cutting 2D-LC focuses on selected regions of interest from the first dimension. Only specific peaks or time windows are transferred to the second dimension for further analysis. This approach allows extended separation time and higher resolution in the second dimension, making it particularly suitable for targeted investigations of known critical compounds. However, because only selected fractions are analyzed, unexpected impurities outside the selected region may remain undetected.<img src="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/KNAUER_Figure_5_2_D_LC_Single_Heart_Cut_Principle_Graphic_by_KNAUER_b1b400784b.jpg" alt="KNAUER: Figure 5 - 2D-LC Single Heart-Cut Principle (Graphic by KNAUER)." srcset="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/thumbnail_KNAUER_Figure_5_2_D_LC_Single_Heart_Cut_Principle_Graphic_by_KNAUER_b1b400784b.jpg 245w," sizes="100vw" width="1276" height="319">In contrast, comprehensive 2D-LC transfers the entire eluate from the first dimension into the second dimension in a continuous manner. This approach provides a complete and detailed representation of the sample, making it ideal for research and development, impurity profiling, and regulatory applications. While more demanding in terms of speed and system performance, comprehensive 2D-LC ensures full sample coverage.<img src="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/KNAUER_Figure_6_Full_Comprehensive_2_D_LC_Principle_Graphic_by_KNAUER_e13da63ef1.jpg" alt="KNAUER: Figure 6 - Full Comprehensive 2D-LC Principle (Graphic by KNAUER)." srcset="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/thumbnail_KNAUER_Figure_6_Full_Comprehensive_2_D_LC_Principle_Graphic_by_KNAUER_e13da63ef1.jpg 245w," sizes="100vw" width="1327" height="292">In practice, hybrid approaches may also be employed, combining targeted heart-cutting with broader comprehensive analysis to balance resolution and coverage.<h4>Practical Considerations for Method Development</h4>Successful implementation of 2D-LC for oligonucleotide analysis requires careful method design.&nbsp;<h5>1. Column selection&nbsp;</h5>It is a critical starting point, as the combination of stationary phases must provide sufficient orthogonality while maintaining compatibility between mobile phases. Typically, the first dimension is optimized for general separation of oligonucleotide size or class, while the second dimension focuses on sequence-specific resolution and compatibility with mass spectrometry.<h5>2. Mobile phase selection&nbsp;</h5>Differences in pH and organic modifiers can significantly influence separation behavior, but incompatibility between dimensions can compromise performance. Therefore, mobile phases must be designed as a coordinated system. The use of volatile buffers, such as ammonium acetate, is often preferred for MS-based detection.<h5>3. Instrumentation&nbsp;</h5>The switching valve must operate reliably under high pressure, with precise timing to ensure accurate fraction transfer. Loop volumes must be carefully selected to avoid overfilling, as excessive loading can lead to sample loss and distortion due to flow profile effects. As a general guideline, loops should not be filled beyond approximately 70% of their capacity.<h5>4. Method development&nbsp;</h5>Typically begins with optimization of the second dimension, as it dictates overall system timing. Fast separations using short columns and steep gradients are often required to maintain sufficient sampling frequency. Once the second dimension is established, the first dimension can be adjusted to achieve optimal resolution and compatibility.<h4>Final Remarks</h4>Two-dimensional liquid chromatography represents a powerful advancement in oligonucleotide analysis, enabling the resolution of complex mixtures that exceed the capabilities of conventional 1D approaches. By combining orthogonal separation mechanisms, 2D-LC reveals structural details and impurity profiles that would otherwise remain hidden.However, achieving optimal performance requires careful balancing of multiple factors, including column chemistry, mobile phase compatibility, system timing, and data acquisition. When properly implemented, 2D-LC provides a robust and highly informative platform for modern oligonucleotide characterization.<blockquote><ul><li>If you're planning to set up or optimize your oligonucleotide workflow, feel free to contact us at <a href="sales@knauer.net">sales@knauer.net</a>. Stay tuned for more exciting insights into the Oligonucleotide world in our “Oligos Made Easy” series.</li><li>For more in-depth discussion or questions, reach out to the author at <a href="kindler@knauer.net">kindler@knauer.net</a></li></ul></blockquote>

Oligos Made Easy - Part 9: 2D-LC

News from LabRulezLCMS Library - Week 14, 2026

<h3>We're excited to announce that <a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/products/999">Clarity Chromatography Software</a> version 10.1.2 is now available!</h3>This minor update expands Clarity’s portfolio with new control modules.<h4>Newly added Instrument control modules</h4><ul><li>Ecom auxiliary modules</li><li>HTA ICF control modules</li><li>Spark pump</li></ul><h4>Updated Instrument control modules</h4><ul><li><a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/companies/168">Ecom</a> HPLC</li><li><a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/companies/328">Knauer</a> HPLC</li><li>Spark ICF control modules</li><li>YoungIn Chromass HPLC</li></ul><blockquote>For a complete list of module changes, see the <a target="_blank" rel="noopener noreferrer" href="https://www.dataapex.com/version_history?product=1">Changelog</a>. For more info about exciting new features directly in Clarity, explore what was introduced in the <a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/article/5622">Clarity 10.1 release highlights</a>.Clarity can control 1000+ instruments; the <a target="_blank" rel="noopener noreferrer" href="https://www.dataapex.com/controls">list can be previewed here</a>.</blockquote><h4>Supported Instruments in Clarity (DataApex) — Quick Guide</h4>Clarity Chromatography Software supports a wide range of chromatographs and peripherals via digital control (DD) and analog acquisition (A/D) using the Colibrick module.<h5>Gas Chromatographs (GC)</h5>Digital integration over LAN/RS232 (many via ICF):<ul><li>Agilent: 5890, 6890, 7890, 8860, 8890, Intuvo 9000</li><li>Shimadzu: GC-2010/2014, Nexis GC-2030</li><li>Thermo Fisher: TRACE 1300/1310</li><li>PerkinElmer: Clarus 400–690</li><li>Also supported: Dani, Ellutia, Young In Chromass, Apix, Fuli, GOW-MAC (selected models)</li></ul><blockquote>Note: Most Agilent 6890–8890 and Intuvo use LAN – ICF.</blockquote><h5>HPLC Systems</h5>Full system/module control:<ul><li>Agilent 1100–1290 Infinity II (ICF)</li><li>Shimadzu LC-10/20 and Nexera LC-40</li><li>PerkinElmer Flexar / NexSAR</li><li>Knauer Azura / BlueShadow / Smartline</li><li>Hitachi Chromaster / Primaide</li><li>Young In Chromass ChroZen series</li><li>Sykam S500–S600, Watrex Streamline, Hanon LC7000–7300</li></ul><h5>HPLC Modules (Pumps, Detectors, Ovens, Fraction Collectors)</h5><ul><li>Pumps: Agilent 1100–1290 families, Knauer, Shimadzu, PerkinElmer, Sykam, SSI, Ecom</li><li>Detectors: VWD/DAD(PDA)/RID/FLD/ELSD across Agilent, Shimadzu, Knauer, PerkinElmer, Young In Chromass, Ecom, etc.</li><li>Column Ovens: Agilent G1316x / G7116x, Shimadzu CTO-20/30/40, Knauer CT2.1, PerkinElmer Flexar</li><li>Fraction Collectors: Knauer FC 6.1, Teledyne FOXY series, Ecom ECF 2096, Waters Fraction Collector II</li></ul><h5>Autosamplers</h5><ul><li>Agilent GC ALS (7673/7683/7693/8697), HPLC ALS modules (G1313A–G7167C)</li><li>Shimadzu AOC-20 / AOC-5000+, SIL-10 / SIL-20 / SIL-40 series</li><li>CTC Analytics PAL, HTA autosamplers, Spark Alias/UHPLC, Knauer AS 6.1L</li><li>PerkinElmer Flexar / NexSAR AS</li></ul><h5>Other Devices</h5><ul><li>GC×GC modulators (e.g., Charles University GC×GC Modulator)</li><li>Valves &amp; switch modules (Agilent, Knauer, Shimadzu, VICI-Valco)</li><li>Degassers, flow monitors, managers, interfaces</li><li>LC-MS / GC-MS: Advion Expression CMS, Microsaic 4500 MiD, Markes BenchTOF (and others)</li><li>Balances (EA extension): Mettler Toledo, Sartorius</li></ul><h5>Analog Acquisition (Colibrick)</h5>A/D support for legacy or analog-output devices:<ul><li>GC: Agilent 5880/4890A, Varian 3xxx/3800/3900, PerkinElmer Clarus series, Thermo TRACE, Shimadzu GC-14/9A, Scion 43x/45x/83xx/85xx, etc.</li><li>HPLC Detectors: Waters 2487/2489, ESA Corona, Hitachi L-series, JASCO FP/UV, Ecom LCD series, many others</li><li>Elemental &amp; Amino Acid Analyzers: Carlo Erba/Costech ECS, Biochrom 20/30+</li></ul>If you are interested in a DEMO VERSION, contact us!

DataApex Clarity Chromatography software 10.1.2 just released!

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Three Webinars and a Roundtable with Experts from Merck KGaA, Darmstadt, Germany, Metrohm &amp; Mettler Toledo
A Week Dedicated to Karl Fischer Titration!
Karl Fischer titration is still the most used, most accurate, and fastest method to determine water content in all kinds of sample matrices. This titration week covers different aspects that need to be considered when determining the water content in products, raw materials, and other substances. Experts from the life science business of Merck KGaA, Darmstadt, Germany, Metrohm AG, and Mettler Toledo will present and discuss different steps of the Karl Fischer titration process and how to improve it. Learnings include:
<ul>
<li>Karl Fischer titration process and tips for how to improve it</li>
<li>Karl Fischer oven method and its advantages</li>
<li>Reference materials to use for titer determination, instrument check, and result verification</li>
<li>Parameters needed to validate a Karl Fischer titration application</li>
<li>News and future aspects of Karl Fischer titration</li>
</ul>
<h3>Importance of CRM-grade Karl Fischer Standards for Accurate KF Titration</h3>
Karl Fischer (KF) titration is one of the most rapid and accurate methods to determine water content in various samples. For accurate KF titration, high quality Certified Reference Materials (CRMs) are essential. In this webinar, we will present key reference material terms, different quality grades of reference materials, and an overview of our portfolio of high-quality CRM-grade KF Standards. By attending this presentation, you will learn:
<ul>
<li>Key terms: Metrological Traceability, Quality Grades, 17025 &amp; 17034 Accreditation</li>
<li>The importance of using the right quality grade of KF standards for your application</li>
<li>An overview of our unmatched portfolio of KF standards</li>
</ul>
<blockquote>
Presenter: Shailly Krishna-Rajusth (Product Manager for Physical &amp; Chemical Property Standards at the life science business of Merck KGaA, Darmstadt, Germany)
</blockquote>

Separation Science: Importance of CRM-grade Karl Fischer Standards for Accurate KF Titration

Importance of CRM-grade Karl Fischer Standards for Accurate KF Titration

Key reference material terms, different quality grades of reference materials, and an overview of our portfolio of high-quality CRM-grade KF Standards.

CRM-grade Karl Fischer Standards for Accurate KF Titration

Three Webinars and a Roundtable with Experts from Merck KGaA, Darmstadt, Germany, Metrohm &amp; Mettler Toledo
A Week Dedicated to Karl Fischer Titration!
Karl Fischer titration is still the most used, most accurate, and fastest method to determine water content in all kinds of sample matrices. This titration week covers different aspects that need to be considered when determining the water content in products, raw materials, and other substances. Experts from the life science business of Merck KGaA, Darmstadt, Germany, Metrohm AG, and Mettler Toledo will present and discuss different steps of the Karl Fischer titration process and how to improve it. Learnings include:
<ul>
<li>Karl Fischer titration process and tips for how to improve it</li>
<li>Karl Fischer oven method and its advantages</li>
<li>Reference materials to use for titer determination, instrument check, and result verification</li>
<li>Parameters needed to validate a Karl Fischer titration application</li>
<li>News and future aspects of Karl Fischer titration</li>
</ul>
<h3>Direct Karl-Fischer Titration or Gas Phase Extraction – A Closer Look</h3>
In this presentation we will compare the direct addition with the gas phase extraction method. For this purpose, various sample types were analyzed by either direct addition, gas phase extraction, or both. Among the tested sample types are powders, oils, and cannabis flower. We will discuss the challenges that such samples represent and highlight the analytical measures that will ensure to obtain the best results possible. By attending this presentation, you will learn about:
<ul>
<li>Direct titration vs. gas phase extraction method</li>
<li>Advantages of the Karl Fischer oven method (gas phase extraction)</li>
<li>Application examples: oils / cannabis flower</li>
</ul>
<blockquote>
Presenter: Thomas Hitz, PhD (Application Development, Automated Karl-Fischer Titration, Mettler Toledo)
</blockquote>

Separation Science: Direct Karl-Fischer Titration or Gas Phase Extraction – A Closer Look

Direct Karl-Fischer Titration or Gas Phase Extraction – A Closer Look

In this presentation we will compare the direct addition with the gas phase extraction method.

Direct Karl-Fischer Titration or Gas Phase Extraction

Three Webinars and a Roundtable with Experts from Merck KGaA, Darmstadt, Germany, Metrohm &amp; Mettler Toledo
A Week Dedicated to Karl Fischer Titration!
Karl Fischer titration is still the most used, most accurate, and fastest method to determine water content in all kinds of sample matrices. This titration week covers different aspects that need to be considered when determining the water content in products, raw materials, and other substances. Experts from the life science business of Merck KGaA, Darmstadt, Germany, Metrohm AG, and Mettler Toledo will present and discuss different steps of the Karl Fischer titration process and how to improve it. Learnings include:
<ul>
<li>Karl Fischer titration process and tips for how to improve it</li>
<li>Karl Fischer oven method and its advantages</li>
<li>Reference materials to use for titer determination, instrument check, and result verification</li>
<li>Parameters needed to validate a Karl Fischer titration application</li>
<li>News and future aspects of Karl Fischer titration</li>
</ul>
<h3>Validation of Parameters for Karl Fischer Titration</h3>
Validation of parameters for a Karl Fischer titration method is an essential prerequisite for laboratories. Performance characteristics of the procedure need to meet the requirements for its intended purpose. You will learn typical analytical performance parameters that should be considered in the validation of the Karl Fischer titration procedures. The objective of this session is to present the requirements for a titration process.
By attending this presentation, you will learn about:
<ul>
<li>General aspects of validation</li>
<li>Required validation parameters</li>
<li>How to verify the suitability and accuracy of a titration method</li>
<li>An overview of the parameters described in ICH (International Council for Harmonization) guidance Q2(R1) and USP general chapter &lt;1225&gt; (selectivity, linearity, accuracy &amp; reproducibility)</li>
</ul>
<blockquote>
Presenter: Angela Schwehn (Laboratory Scientist in the R&amp;D Department of the life science business of Merck KGaA, Darmstadt, Germany)
</blockquote>

Separation Science: Validation of Parameters for Karl Fischer Titration

Validation of Parameters for Karl Fischer Titration

The objective of this session is to present the requirements for a titration process.

Three Webinars and a Roundtable with Experts from Merck KGaA, Darmstadt, Germany, Metrohm &amp; Mettler Toledo
A Week Dedicated to Karl Fischer Titration!
Karl Fischer titration is still the most used, most accurate, and fastest method to determine water content in all kinds of sample matrices. This titration week covers different aspects that need to be considered when determining the water content in products, raw materials, and other substances. Experts from the life science business of Merck KGaA, Darmstadt, Germany, Metrohm AG, and Mettler Toledo will present and discuss different steps of the Karl Fischer titration process and how to improve it. Learnings include:
<ul>
<li>Karl Fischer titration process and tips for how to improve it</li>
<li>Karl Fischer oven method and its advantages</li>
<li>Reference materials to use for titer determination, instrument check, and result verification</li>
<li>Parameters needed to validate a Karl Fischer titration application</li>
<li>News and future aspects of Karl Fischer titration</li>
</ul>
<h3>Live Roundtable Expert Discussion (Karl Fisher Titration Week)</h3>
Together with our partners Metrohm and Mettler Toledo, Merck KGaA, Darmstadt, Germany will discuss the trends and key aspects of Karl Fischer titration. We will come back on some questions asked during the earlier three webinars, as well as answering new questions during the roundtable discussion. 
This live event will be moderated by Separation Science and participants will include Bettina Straub-Jubb, Dr. Shailly Krishna Rajusth, and Angela Schwehn from Merck Life Science KGaA, Darmstadt, Germany, Dr. Leila Mosberger from Mettler-Toledo, and Michael Margreth from Metrohm AG.

Live Roundtable Expert Discussion (Karl Fisher Titration Week)

We will discuss the trends and key aspects of Karl Fischer titration.

Live Roundtable Expert Discussion (Karl Fisher Titration)

Data independent acquisition (DIA) allows for suspect screening in complex matrices with retroactive analysis capabilities important to emerging compounds of interest in food safety. However, with increased analyte scope, the review of questionable identifications can create an analysis burden requiring timely review and reinjections.
Here we present a robust DIA methodology with a novel prototype LC/Q-TOF, leveraging a combination of high-resolution, extended dynamic range, stable accurate mass, and isotopic fidelity with coeluting fragmentation ions for identification, analyzed with MH Quantitative Analysis 12.1 and LC Screener Tool.
Over 200 analytes were identified from a spiked broccoli matrix prepared with QuEChERS, resulting in good calibration curve linearity and low area %RSDs. Fused, is the Intelligent Reflex Workflow used to automatically evaluate and reinject samples in which results were flagged as questionable and/or identified, via targeted MS/MS utilizing the available m/z and RT information. Transferring questionable results into confirmed identifications by acquiring targeted data for present compounds. Leveraging the combination of DIA and automatically enabled targeted MS/MS reinjections via Intelligent Reflex allows for broad screening analysis and confident identification confirmation of analytes.
<blockquote>
Presenter: Cathyrin Simmermaker (LC/MS Applications Engineer, Agilent Technologies, Inc.)
</blockquote>
<blockquote>
Cate Simmermaker is an LC/MS Applications Engineer with Agilent Technologies working primarily with high-resolution mass spectrometry in small molecule analysis. Cate has worked in applications including metabolomics, food safety, and forensics on Q-TOF and QQQ platforms. She is currently pursuing her PhD in Chemistry from the University of the Pacific and received a Master’s in Biochemistry from San Francisco State University.
</blockquote>
<blockquote>
Presenter: Christian Klein, PhD (Product Manager LC/Q-TOF, Agilent Technologies, Inc.)
</blockquote>
<blockquote>
With an education in chemistry, Chris Klein received his PhD in 2005 at the Max-Planck-Institute for Biochemistry in Martinsried, Germany, working on membrane proteins and mass spectrometry. After postdoctoral research and three years as a demo chemist and product specialist with Bruker Daltonics, Chris joined the Agilent LC/Q-TOF research and development team in 2010. Since 2018, he has been the LC/Q-TOF and IM-QTOF product manager.
</blockquote>

Agilent Technologies: From Screening to Confirmation: A Comprehensive Food Safety Analysis from Initial DIA Screening to Automatic Reinjection for Suspect Confirmation

From Screening to Confirmation: A Comprehensive Food Safety Analysis from Initial DIA Screening to Automatic Reinjection for Suspect Confirmation

Here we present a robust DIA methodology with a novel prototype LC/Q-TOF.

From Screening to Confirmation: Food Safety Analysis

First of all, Lakshmanan Deenadayalan will present a sensitive method for the quantification of formoterol – a drug used to treat asthma and COPD – in human plasma.
Then, Eshani Nandita will introduce a sensitive multiplexed quantification of two commonly co-administered mAbs using the ZenoTOF 7600 system with SCIEX OS software.
Finally, Ian Moore will show us how to achieve increased sensitivity for quantification in complex matrices using the SCIEX 7500 system.
At the end of the presentations, you will be able to interact with the speakers and ask any questions.
Attend this webinar to:
<ul>
<li>Meet front-end technology that improves your quantitative mass spectrometry methods</li>
<li>Achieve ultra-low quantification levels for compounds in human plasma using a simple liquid-liquid extraction sample preparation</li>
<li>Reach LLOQs of 0.15 µg/mL for the quantification of monoclonal antibodies in human serum with a QTOF instrument</li>
<li>Explore lower dosage, higher efficacy compounds and improve the efficiency of bioanalysis in negative mode</li>
</ul>
<blockquote>
Presenter: Ian Moore (Senior Technical Product Manager, Nominal Mass SCIEX)
</blockquote>
<blockquote>
Presenter: Lakshmanan Deenadayalan (Application Support Specialist, Indian Innovation Center (IIC) SCIEX)
</blockquote>
<blockquote>
Presenter: Eshani Nandita (Global Biopharma Quant Tech Marketing Lead, SCIEX)
</blockquote>

Pixabay/Gerd Altmann: Inside the Tech Note: The Story Behind the Data

Inside the Tech Note: The Story Behind the Data

In this webinar, hear three LC/MS Tech Note authors present their work and explain the genesis of their research.

Robustness testing is a measure of a method’s ability to remain unaffected by the minor changes of chromatographic parameters. The robustness multivariable design of experiments (DoE) can be used to study the effect of individual parameters and their interaction on the chromatographic resolution between peaks.
The Empower Sample Set Generator (SSG) can be used to automatically create a sample set method, which includes instrument methods, and method sets required to perform the entire study in one run. The Effect Plots generated by the Method Validation Manager clearly show which parameters have the greatest effect on method performance and provides knowledge for establishing a set of controls to assure a method meets the expected criteria during routine use.
Key points of discussion will be:
<ul>
<li>Using the Sample Set Generator with the Method Validation Manager option to easily create Sample Sets needed for the Robustness test
</li>
<li>The Robustness multivariable design of Experiments (DoE) is easily created to study the effect of parameters and their interaction on chromatographic resolution between peaks
</li>
<li>The Effect Plot in the Report Publisher can be used to clearly shows which parameters have the greatest effect on method performance
</li>
</ul>
Join us for a live discussion as we explore Empower 3.8.0. As always, don’t forget to bring your questions for this exclusive opportunity to ask our experts anything about Empower Software
<blockquote>
Presenter: Neil Lander (Principal Product Manager, Waters Corporation)
</blockquote>
<blockquote>
Neil has worked for Waters Corporation since 1994 and has held a variety of positions including Technical Support Specialist, Technical Trainer, Manager Customer Education and currently, Principal Marketing Manager focusing on Informatics solutions. He provides internal support for Empower Chromatography Data System and is focused on developing laboratory software solutions that help organizations achieve their scientific and operational objectives. Prior to joining Waters, he was an analytical chemist where he gained experience in both the cosmetics and specialty chemical industries.
</blockquote>

Waters Corporation: Smart Method Development using the Empower Method Validation Manager and the Sample Set Generator

Smart Method Development using the Empower Method Validation Manager and the Sample Set Generator

Using the Sample Set Generator with the Method Validation Manager option to easily create Sample Sets needed for the Robustness test.

Smart Method Development using the Empower Method Validation

There are a variety of sample preparation methods used for the analysis of drugs of abuse in physiological sample matrices such as blood, urine, oral fluid.
These include simple ‘dilute and shoot’, protein removal, phospholipid removal, liquid -liquid and solid phase extraction as well as many others.
With each method there is variability in the number of steps involved, specificity, and the suitability to automation.
Choosing the most suitable sample preparation technique can depend on the type of analytical challenge being addressed, for example certain methods can be used to screen for a large number of compounds and others can be selected to enhance sensitivity for a particular compound or class of compounds. 
In this session, a number of sample preparation methods will be discussed in terms of their strengths and limitations and suitability for particular forensic toxicology applications.
This webinar will prepare the audience for the third session in the Forensic Toxicology Summer Webinar series where the use of a simple generic protocol is used to analyse a wide range of drugs of abuse in whole blood to support driving under the influence of drugs investigations.
Learning objectives:
<ul>
<li>Uncover an overview of selective and generic methods for biological fluids</li>
<li>How these sample preparation techniques can be applied to your most complex toxicological samples</li>
</ul>
<blockquote>
Presenter: Jonathan Danaceau, Ph.D. (Principal Scientist, Waters Corporation)
</blockquote>
<blockquote>
Dr. Jonathan Danaceau, Ph.D. is a Principal Scientist with Waters. He received his B.S. in Biology from Allegheny College and a Ph.D. in Neuroscience from the University of Utah (Salt Lake City, UT). Jon came to Waters in 2011 after spending several years in the forensic toxicology and sports doping fields. Since joining Waters, he has focused on sample preparation and LC/MS/MS solutions for forensic, clinical research, and other bioanalytical applications. He is an expert in sample preparation and bioanalytical method development and has published several papers in these fields. Jon is currently with the Sample Preparation and Automation group within the Consumables and Lab Automation division at Waters.
</blockquote>

Waters Corporation: Sample Preparation Strategies for Forensic Toxicology Testing

Sample Preparation Strategies for Forensic Toxicology Testing

A number of sample prep methods will be discussed in terms of their strengths and limitations and suitability for particular forensic toxicology apps..

Sample Preparation Strategies for Forensic Toxicology

As a continuation of our Chromatography of Bioseparations 101 series, we bring to you 2 NEW SESSIONS in our Chromatography of Bioseparations 102 webinar series! Join us live for our new parts 6 and 7. If you haven&#39;t had an opportunity to register for Parts 1-5, feel free to regster and listen to them on-demand now.
These new webinars will focus on the analytical separation of biomolecules such as peptides and amino acids.
Reverse-phase chromatography is the predominant technique used for the analyses and or purification of chemically synthesized peptides or peptides generated by the enzymatic digestion of proteins (i.e., peptide mapping). It is advantageous to have a simplified step-by-step approach to develop a method that can be applied to chemically synthesized peptides or protein digests.
Amino acid analysis is required in several different areas of research and is a fundamental tool in various product analytical activities. Due to the varied properties of amino acids (i.e., acidic, basic, etc.), the separation and detection represents an analytical challenge and depending on throughput, sample preparation can be laborious. 
Who should attend?
<ul>
<li>Those interested in a refresher on foundational principles and tips of reverse phase separations of peptides.</li>
<li>Those interested in how to select columns and mobile phases that are optimal for peptide and amino acid workflows.</li>
<li>Those looking for an overview of industry advancements and new offerings.</li>
</ul>
<h3>PART 6 | June 20th, 2023 | 10:30AM – 12:00PM EDT</h3>
<h3>PRINCIPLES AND PRACTICAL STEPS IN DEVELOPING REVERSE-PHASE PEPTIDE SEPARATIONS</h3>
<ul>
<li>Learn how column chemistry and mobile phase affects peptide separation and selectivity.</li>
<li>Review the factors that synergistically work to obtain desired resolution of peptides contained in a complex mixture.</li>
</ul>
<blockquote>
Presenter: Bill Warren (Consulting Product Manager – Consumables and Lab Automation, Waters Corporation)
</blockquote>
<blockquote>
Bill received his MS in Microbiology and Immunology from the University of Louisville School of Medicine. In 1986, joined Waters Corporation as an senior applications chemist involved with the research and development of HPLC, FPLC, and Capillary Electrophoresis applications for the analysis and purification of proteins, peptides, and nucleic acids. Following four years as a field based technical sales representative for Waters, Bill enjoyed a tenure as Manager, Core Product Evaluation Laboratory within the Waters Worldwide Marketing Organization. Bill currently is a Principal Bioseparation Columns Product Manager whose responsibilities involve assisting in the development and commercialization of innovative reagents and chromatography-based technologies for the separation, purification, and characterization of proteins and peptides.
</blockquote>

Waters Corporation: PRINCIPLES AND PRACTICAL STEPS IN DEVELOPING REVERSE-PHASE PEPTIDE SEPARATIONS

PRINCIPLES AND PRACTICAL STEPS IN DEVELOPING REVERSE-PHASE PEPTIDE SEPARATIONS

Learn how column chemistry and mobile phase affects peptide separation and selectivity. Review the factors that synergistically work to obtain resolution.

DEVELOPING REVERSE-PHASE PEPTIDE SEPARATIONS

Purification is critical to many scientific workflows. Compounds of interest are rarely found to be the right purity in their native state. Join Waters and Virscidian for this 6-part webinar series to learn the basics of purification and application of purification principles for more advanced workflows.
<h3>PART 3 | Things to Consider When Configuring Purification Systems and Informatics</h3>
Preparative chromatography is a separation technique where one or more peaks are collected for use in another experiment or process.
In this module, we will discuss the various hardware options available, from pumps and injectors through detection techniques to fraction collection. We’ll also touch on some of the software options available and look at different schemes for triggering fraction collection. 
Learning Objectives: 
<ul>
<li>Understand various instrument configuration and the impact on purification</li>
<li>Optimize system performance</li>
</ul>
<blockquote>
Presenter: Darcy Shave (Senior Product Manager, Waters Corporation)
</blockquote>
<blockquote>
Darcy Shave is currently aprincipalproduct manager in the Integrated Systems group atWaters Corporation (Milford, MA, USA), responsible for the purification and hardwareautomation products. Darcy has been with Waters for over 20 years, starting as anapplications chemist in the purification group. From there, he moved into a variety ofbusiness development roles before becoming a product manager for purification softwareabout 10 years ago and more recently for the entire purification portfolio.​
</blockquote>
<blockquote>
He has a degree in Agricultural Chemistry from the University of Saskatchewan and workedin value added processing of natural products before joining Waters.
</blockquote>

Waters Corporation: Things to Consider When Configuring Purification Systems and Informatics

Things to Consider When Configuring Purification Systems and Informatics

We will discuss the various hardware options available, from pumps and injectors through detection techniques to fraction collection.