This technical note explores the advantages of using relative standard error (RSE), weighted least squares (WLS) approximations, and inverted calibrations to enhance calibration quality in chromatography. By optimizing calibration curves through quadratic fitting, offset adjustments, and suitable weighting factors (such as 1/Amount²), the calibration process can be significantly improved, resulting in lower RSEs and better overall calibration quality. Incorporating RSE as an evaluation tool offers a deeper understanding of calibration reliability.

HPAE-PAD is a well-established technique for separating and quantifying a wide range of carbohydrates, providing high resolution separation and high sensitivity. This document discusses the basics of the separation and detection mechanisms and limitations of HPAE-PAD, good practices and techniques for successful analyses.

The proper preparation and use of eluents are the most crucial parts for success in carbohydrate analysis with ion chromatography with PAD detection. This document describes purity criteria for reagents, proper eluent storage, proper use, identification of contaminated eluent and consequences.

The presented 2D-LC-MS system facilitates the straightforward analysis of mAb producing cell culture samples for the upstream monitoring of titer, charge variants, and size variants of the intact antibody. While the first dimension purifies the mAb from matrix components by Protein A affinity chromatography, selectable second dimension mechanisms enable the assessment of the aggregation level by SEC or the abundance monitoring of glycoforms and other variants by SCX.

Exploring the potential of a novel, intelligent tandem LC-MS workflow with near 100% MS utilization from deep-dive and high-throughput proteomics.

Demonstrating a tandem LC-MS configuration for high-throughput proteomics and peptide quantification with near 100% MS utilization.

To demonstrate a targeted LC-UV-SQMS-based peptide monitoring workflow for QC labs, enabling increased productivity in sample preparation, data analysis, and reporting.

The Thermofisher Scientific Vanquish™ Flex UHPLC system was used to analyze Tocilizumab Charge variant analysis using Propac 3R SCX Column and CX-1 Buffer

The Thermofisher Scientific Vanquish™ Flex UHPLC system was applied for the analysis of Charge related impurities in obinutuzumab using Propac 3R Column and CX-1 buffer

Tocilizumab Oxidized Impurity analysis using Thermo Fisher Mabpac HIC-20 Column. Thermo Scientific™ Ultimate 3000™ UHPLC Bioinert System.

The Thermo Scientific™ Ultimate3000™ Biointert UHPLC system was applied for the aggregate analysis of Ustekinumab using Mabpac SEC-1 column.

Enable fast and automated mass confirmation for oligonucleotides, streamlining data acquisition, processing, and reporting of LC-MS based intact mass analysis with compliance-ready Chromeleon CDS. Quick and confident confirmation of modified oligonucleotides on an Thermo Scientific™ ISQ™ EM Single Quadrupole Mass Spectrometer

Using the Vanquish Analytical LC Purification System for the semi-preparative reversed-phase liquid chromatographic purification of oligonucleotides

Multiple heart-cut 2D-LC-MS is an efficient approach for charge variant analysis of antibodies by enabling direct MS analysis of first dimension ion-exchange fractions after desalting in the second dimension. Intact mass analysis facilitates identification and monitoring of mAb charge variant patterns. Annotations of major and minor charge variant peaks could be proposed. Enrichment of one fraction was used to improve the MS signal intensity.

Ion-exchange high-performance liquid chromatography (IEX-HPLC) analysis were performed to quantitate degraded impurities of Teriparatide with a fluorescence detector. The analysis was performed on a Thermo Scientific MAbPac SCX-10 RS HPLC column. The separation results were satisfactory.

The Thermo Scientific Dionex Ultimate 3000 Bio LC System is applied for the impurity analysis of Teriparatide formulation sample. Analysis is performed on Mabpac SEC -1

The Thermo Scientific Dionex Ultimate 3000 LC System is applied for the impurity analysis of Streptokinase formulation sample. Analysis is performed on Acclaim 300 C18 column.

The Thermo Scientific Dionex Ultimate 3000 Bio LC System is applied for the impurity analysis of Ranibizumab formulation sample. Analysis is performed on Mabpac SEC-1 column.

The Thermo Scientific Dionex Ultimate 3000 Bio LC System is applied for the impurity analysis of Nivolizumab formulation sample. Analysis is performed on MAbpac SEC-1 300A° pore size column.

The Thermo Scientific Dionex Ultimate 3000 Bio LC System is applied for the charge related impurity analysis of monoclonal antibody IGG4 formulation sample. Analysis is performed on propac SAX Column

The Thermo Scientific Dionex ultimate 3000 Bio LC system is Applied for the Aggregate impurities analysis of Filgrastim. The analysis was performed on Mabpac SEC-1 using the IP monograph method. IP has revised the resolution criteira from 2.0 to 3.0. The critical resolution of Aggregate and monomer was more than 3.0 .

In this application note, we demonstrate a workflow combining the chemical release of O-linked glycans, followed by sample cleanup and analysis by HPAE-PAD hyphenated to a Q Exactive HF mass spectrometer. The Dionex CarboPac PA300-4μm column uses a novel column technology that enables the simultaneous separation of neutral and charged glycans without the need of derivatization. High resolution accurate mass MS data and tandem MS/MS spectra with diagnostic fragment ions provide highly reliable structural annotations of heterogenous glycans.

Demonstrate the performance of the multi-draw functionality using the Thermo Scientific™ Vanquish™ Neo UHPLC system for large volume injections in bottom-up proteomics experiments within the trap-and-elute workflow.

This application note describes a method for nucleotide sugar determination using anion-exchange chromatography. This method uses a Thermo Scientific™ Dionex™ CarboPac™ PA1 column and manually prepared eluents for separation. Using a this column, a 34 min method that resolved seven analytes was designed.

In this paper, a simple and rapid online determination method for molecular weight of DNA primers by ULTRA-performance liquid chromatography (UPLC-VANQUish DuO-LTQ XL) was developed based on the Vanquish Duo Flex UPLC-PLATFORM. This series of methods will provide guidance for the further development of the dual column alternating high-throughput intelligent analysis of DNA primer samples from Vanquish Duo and LTQ XL.

A method was developed for the determination of liposomes in mRNA vaccines using a Vanquish Flex ULTRA-high Performance liquid chromatography combined with a mass universal detector (CAD) electromist detector. This method has high sensitivity, operation detection and wide linear range, and can be used to determine the content of liposome in vaccines.

Because PEG has no uv absorption, it is important to select a universal detector with high sensitivity and high repeatability for quality control of free PEG in polypeptide drugs. CAD has been successfully detected in a variety of different molecular weight PEG. In this paper, 20000 molecular weight PEG detection was tried for the first time, and the performance of ELSD detection was preliminarily compared.

In this paper, a simple and efficient HPLC method was established by using Vanquish Core high performance liquid chromatography (Vanquish Core) combined with general quality detector-Charged Aerosol Detection(CAD). It can meet the needs of simultaneous determination of Poloxam 188 and Tween 80 in adeno-associated virus (AAV) vector preparation. The method has good stability and high sensitivity. The wide linear range is suitable for the determination of different concentrations of samples, and can be used as an effective means for the control of excipient residues in drugs such as Poloxam 188

High-throughput analysis of oligonucleotides using a single quadrupole mass spectrometer for quality control

The Thermo Scientific™ Vanquish™ Duo for Dual LC combined with the Method Scouting Kit offers a valuable solution for determining the most promising chromatographic conditions in a time-effective manner. With the Vanquish Duo system, two independent chromatographic chemistries (IEX and IP-RP) are scouted on the same system at the same time, which enables faster method development and requires less instrumentation. In this work, a large number of mobile phase conditions are tested with minimal preparation work.

This work describes an HPAE-PAD method for the determination of ribitol in Hib capsular polysaccharide acid hydrolysis samples. Separation of ribitol from TFA acid hydrolysis products was achieved under isocratic elution conditions using a Thermo Scientific™ Dionex™ CarboPac™ MA1 column. The method proposed here was validated for performance with respect to linearity, precision, sensitivity, and accuracy. The acid hydrolysis samples were also tested using a palladium hydrogen reference electrode.

KDO (3-deoxy-D-manno-oct-2-ulosonic acid) is an eight-carbon sugar mostly confined to Gram-negative bacteria. KDO can be used to quantify the amount of a known lipopolysaccharide (LPS) (the moles of KDO per mole of liposaccharide must be known). The hydrolysis conditions were optimized for the release of KDO from an LPS and a method for its quantitation has been developed using HPAE-PAD. In this application note, we develop, optimize, and validate a HPAE-PAD method for the determination of KDO.

Tromethamine is commonly used as a buffering agent, alkalizer, and emulsifying agent in pharmaceutical and cosmetic preparations, and as a counterion for acidic drug substances. This work describes an IC method that uses a Thermo Scientific™ Dionex™ IonPac™ CS20 cation exchange column, electrolytically generated MSA eluent, and suppressed conductivity detection to determine tromethamine in pharmaceutical formulations.

The application note describes an efficient approach for identity confirmation of protein-based impurities of a therapeutic protein by multi heart-cut 2D-LC-MS in a top-down experiment. In addition the Thermo Scientific™ Simple Switch™ technology enables the instrument to be used in multi heart-cut 2D-LC-MS, 1D-LC-MS, or Dual 1D-LC mode for the most flexible equipment and lab space utilization and maximum MS productivity.

We demonstrate a workflow combining chemical release of O-linked glycans, followed by sample cleanup and analysis by HPAE-PAD hyphenated to a Q Exactive HF Hybrid Quadrupole-Orbitrap MS. A recently introduced Dionex CarboPac PA300-4μm column, enables simultaneous separation of neutral and charged glycans without the need for derivatization. High-resolution MS data and MS/MS spectra with diagnostic fragment ions provide reliable structural annotations of heterogeneous glycans. We applied HPAE-PAD/MS to analyze O-glycan structures released from four different glycoproteins.

This record has QAR conditions and results for the 2 mm Dionex IonPac CS19-4µm column. The column must be pre-conditioned offline with acetonitrile and methanesulfonic acid (MSA). See instructions in the column manual (attached). The column is optimized for small, hydrophilic amines such as ethanolamines, methylamines, ethylamines, as well as the biogenic amines.

This application note demonstrates that the AU180 method for sialic acid determination of glycoproteins could be successfully modified for use with a Dionex CarboPac PA20-1 mm column and HPAE-PAD in dual eluent generation cartridge mode.

Single-use technologies (SUTs), in particular single use bioreactors (SUBs), represent an important improvement in biopharmaceutical manufacturing due to reduced requirements for cleaning and sterilization while providing increased sterility assurance, reduced manufacturing turnaround times, and the elimination of cleaning validation and its associated costs. The present work explored the performance of ASE as an extraction technique for characterization of the plastic films (inner layers) from SUBs.

Thermo scientific BioLC ultimate 3000RS instrument is used for the analysis of different form of plasmid DNA. The analysis is performed on high resolution anion exchange DNAPac PA200RS column.The result obtain shows well separation of linear and supercoiled form of plasmid DNA.

This technical note (TN) describes how to develop an ion chromatography method to determine an amine in pharmaceutical samples. Two methods were developed in this TN. Method choice depends on the nature of the sample. This record is for the method that uses the Dionex IonPac CS19 column .

This technical note (TN) describes how to develop an ion chromatography method to determine an amine in pharmaceutical samples. Two methods were developed in this TN. Method choice depends on the nature of the sample. This record is for the method that uses the Dionex IonPac CS16 column .

The thermo Scientific ultimate 3000 BioLc system is applied for impurities / isoform analysis of Pertuzumab. The analysis was performed on a thermo scientific MAbPac HIC 10 column.

A method was developed for the determination of nitrite in pharmaceuticals by coupling IC with UV absorbance detection. The LOD of nitrite in a pharmaceutical sample is 0.918 ppm (μg/g API). The method is accurate and precise due to the high reproducibility of the Reagent-Free ion chromatography system. This method should be applicable to the determination of nitrite throughout the manufacturing process of a drug product to assess the likelihood of nitrosamine formation.

An HPLC-based approach to monitor the characteristics of polysorbate 80 samples that provides operational simplicity while generating a high degree of information is proposed. The reversed-phase method is highly optimized to ensure sufficient separation of different compound classes within a reasonable run time to permit the fingerprinting of polysorbate 80 samples and detect variability between different suppliers, grades, and production batches.

The Thermo Scientific Dionex UltiMate 3000 LC Bio-LC system is applied for the Ion Exchange analysis of ranibizumab sample. The separation was performed on a Thermo Scientific MAbPac SCX-10 with a UV detection at 280 nm.

The Thermo Scientific Dionex Ultimate 3000 system is applied for the peptide mapping analysis of an Pertuzumab (IgG1). The separation was performed on a Thermo Scientific Acclaim RSLC 120 C18 column by using a Thermo Scientific SMART digest kit at UV detection of 214 nm. The SMART Digest kit provides significant improvements in reproducibility, which results in fewer sample failures, higher throughput and the ability to more easily interrogate data..

In this application note, the nitrite determination described in the USP monograph was evaluated with a Thermo Scientific Dionex IonPac AS15 column using an IC system. The method and conditions were exactly as described in the USP Dalteparin Sodium monograph. Key performance parameters were evaluated including the system suitability separation, linearity, limits of detection, accuracy, and robustness. One Dalteparin Sodium sample was analyzed. The percentage of nitrite result was compared with the USP acceptance criterium.

The Thermo Scientific Dionex UltiMate 3000 LC Bio-LC system is applied for the Ion Exchange analysis of teriperatide. The separation was performed on a Thermo Scientific MAbPac SCX-10 and fluorescence detector.

Analysis of protein aggregation and fragment of pertuzumab by size-exclusion chromatography, showing the universal applicability of the Thermo Scientific MAbPac SEC-1 column for aggregate and fragment analysis. Aggregate analysis is major CQA for protein based drugs and requires tight monitoring and control.

Combining the Thermo Scientific™ SMART Digest™ magnetic kits with the Thermo Scientific™ KingFisher™ Duo Prime purification system provides an automated approach to protein digest method optimization. A robust and reproducible, automated digestion time-course protocol to determine the optimal digest time for a biotherapeutic during development of a peptide mapping method.