Catecholamine metabolites vanillylmandelic acid (VMA), homovanillic acid (HVA) and 5-hydroxyindoleacetic acid (5 HIAA) are biomarkers for neuroblastoma and catecholamine-secreting tumors. This poster presents optimization of the method development process to maximize analyte sensitivity in the extraction and quantitation of these metabolites from plasma. Sample preparation protocols using solid phase extraction (EVOLUTE EXPRESS ABN and AX) and supported liquid extraction (ISOLUTE SLE+) are presented. MSACL NA 2018 Palm Springs, CA
The use of LC/MS analysis in the clinical lab has increased exponentially over the last 10 years. Modern mass spectrometers are extremely sensitive allowing low level detection of many target analytes. However, this sensitivity can come at a price, in that levels of contamination not previously detected with less sensitive instruments can now have larger impact on analysis. The complexity of common matrices such as plasma/serum and urine while presenting different challenges can have a marked influence on method performance. As a result sample preparation is an extremely important part of the process in order to provide robust methods. This seminar focuses on some of the method development challenges our lab faced when looking at two clinical assays: Endogenous steroid hormone extraction from serum, and catecholamine extraction from plasma and urine. Particular emphasis is placed on the various sample preparation options we looked at for the extraction of these analytes. During optimization of the extraction process we investigated analyte recovery, co-extractable matrix components, HPLC column degradation, calibration curve performance and limits of quantitation. MSACL US 2018 Breakfast Seminar
This poster discusses the impact of optimization of various parts of the method development process to maximize assay performance while delivering a combined assay for the analysis of urinary catecholamines and metanephrines. The extraction method using EVOLUTE EXPRESS WCX 96-well plates is successfully automated using the Biotage® Extrahera™ Automated Sample Preparation Platform. MSACL 2017, Palm Springs ASMS 2017, Indianapolis
This application note describes a mixed-mode weak cation exchange SPE protocol for the extraction of three catecholamines (epinephrine, norepinephrine and dopamine) and three metanephrine metabolites (metanephrine, normetanephrine and 3-methoxytyramine) from urine prior to LC-MS/MS detection. Included in this application note are the detailed settings for implementing the method on the Biotage® Extrahera(TM) automation system.
This poster investigates various parts of the SPE method development process to develop a highly sensitive LC-MS/MS method for analysis of catecholamines (epinephrine, norepinephrine, dopamine). A number of steps are optimized to improve sensitivity of the analytes. This includes reducing the buffer strength in the mobile phase, reducing the buffer strength of washes, careful pH control throughout the extraction, avoiding an evaporation step and incorporating IPA in the reconstitution solvent.
This poster describes a method for the simultaneous extraction (using EVOLUTE EXPRESS WCX) and analysis of plasma catecholamines and metanephrines. A highly sensitive, linear and rugged assay was developed, and automated using the Biotage Extrahera sample preparation processing system. MSACL EU 2016
This poster examines various factors in the development and optimisation of a SPE-LC-MS/MS method for extraction of catecholamines from plasma. The final method utilizes EVOLUTE EXPRESS WCX plates in either standard SPE or fast Load-wash-elute modes, and incorporates a series of wash steps to optimise analytical sensitivity. Low elution volumes mean no evaporation step is required, and LLOQ is 20mg/mL or lower for each of the analytes.
This application note describes a mixed-mode weak cation exchange solid phase extraction protocol using EVOLUTE® EXPRESS WCX 96-well plates for the extraction of three catecholamines (epinephrine, norepinephrine and dopamine) from human plasma prior to LC-MS/MS detection, which allows low level quantification of the analytes.
This report details a “load, wash, elute” weak cation exchange solid phase extraction procedure amenable to both plasma and urine samples. The extracts are subsequently injected into an LC-MS/MS system. The preliminary sample preparation method was developed at the Biotage US Applications lab (Charlotte, NC).The method was then transferred to Ionics (Bolton, ON, Canada) to facilitate the nmole/L measurements of the selected biomarkers by laminar flow tandem mass spectrometry. The SPE-LC-ESIMS/ MS method parameters were first optimized using pooled mixed gender plasma. A set of patient samples (n=32) was later supplied by the Mayo Clinic (Rochester, MN) that had previously been analyzed by a validated referee method. The population represented measured values across a range of clinical relevance. AACC 2014
This application note describes a method of extraction for metanephrine and normetanephrine from pooled plasma using EVOLUTE® EXPRESS WCX 96-well SPE plates. A new solid phase extraction technology has been developed to reduce the time associated with sample preparation methods by eliminating traditional steps in the extraction workflow. Leveraging this technology, a method protocol was developed using a load, wash and elute approach. This method was optimized for pooled plasma.
This application note describes a method of extraction for metanephrine and normetanephrine from synthetic urine using EVOLUTE® EXPRESS WCX 96-well SPE plates. A new solid phase extraction technology has been developed to reduce the time associated with sample preparation methods by eliminating steps in the extraction workflow. Leveraging this technology, a method protocol was developed using a load, wash and elute approach. This method was optimized for synthetic urine.
This method details the use of a non-polar retention mechanism for the extraction of catecholamines (noradrenaline, adrenaline, dopamine) from urine. The polar analytes form a non-polar complex with phenylboronic acid and are retained on the non-polar MFC18 column. Typical recoveries are >85%. ISOLUTE, Catecholamines, dopind, sports drug testing,