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

96-well plate Catecholamines Clinical English LC-MS/MS Plasma Serum Steroid Hormones Steroid metabolite Urine

This application note demonstrates that the Biotage(R) Horizon 4790 using Atlantic C18 disks can effectively extract hormone compounds from 1-L water samples in approx 40 mins.

11-ketotestosterone 17-alpha-estradiol 17-alpha-ethynylestradiol 17-beta-estradiol 4790 Androgens Androsterone Application Notes Diethylstilbestrol Dihydrotestosterone (DHT) Disk Endocrine disrupter English Environmental Epitestosterone Estradiol Estriol Estrogens Estrone GC-MS Hormones Mestranol Norethindrone Progesterone Steroid Hormones Steroid metabolite Surface water Testosterone Water

Whether you need targeted methods for analytes such as Vitamin D metabolites in serum, or methods suitable for extraction of a wide panel of drugs and metabolites from urine, sample preparation before analysis is essential. This compendium highlights a selection of clinical sample preparation applications from Biotage.

Alcohol metabolite Application Notes Biomarker Brochures Catecholamines Clinical English Pain Management Peptides Product Notes Steroid Hormones Steroid metabolite Thyroid Hormone Tobacco Specific Nitrosamines (TSNAs) Vitamin D Vitamins Water Soluble Vitamins

This poster compares sample preparation options (solid phase extraction using EVOLUTE EXPRESS ABN, and supported liquid extraction using ISOLUTE SLE+) for the extraction of a panel of endogenous steroids from serum. MSACL NA 2018, Palm Springs, CA ASMS 2018, San Diego, CA

96-well plate Clinical English LC-MS/MS Posters Serum Steroid Hormones Steroid metabolite

This poster compares sample preparation options for the extraction of a panel of endogenous steroids from serum. LC-MS/MS parameters were investigated for increased sensitivity: MRM transitions, chromatography and mobile phase additives for use with positive and negative ionisation modes. Particular emphasis was placed on the sample preparation to provide high reproducible recoveries whilst minimizing matrix effects and co-extracted materials such as proteins and phospholipids. Solid phase extraction was compared to supported liquid extraction in terms of recoveries, ion suppression, phospholipid content, calibration curve performance and overall sensitivity. MSACL EU 2018

96-well plate Clinical English LC-MS/MS Posters Steroid Hormones Steroid metabolite

This poster compares sample preparation options for the extraction of a panel of endogenous steroids from serum. EBF 2019

96-well plate Clinical English LC-MS/MS Posters Serum Steroid Hormones Steroid metabolite

This application note describes the extraction of a panel of 18 steroid hormones from horse hair using ISOLUTE® SLE+. The simple sample preparation procedure delivers clean extracts and analyte recoveries greater than 80% with RSDs lower than 5% for most analytes. Linearity of greater than 0.999 is achieved for all analytes in the range 0.5-500 pg/mg of hair.

11-deoxycorticosterone 11-deoxycortisol 17-alpha-hydroxyprogesterone 21-deoxycortisol 96-well plate Aldosterone Anabolic steroids Androstenedione Application Notes Corticosterone Cortisone DHEA (dehydroepiandrosterone Dehydroepiandrosterone (DHEA) Dihydrotestosterone (DHT) English Estradiol Estrone Forensic Hair Horse Hair LC-MS/MS Lysera Progesterone Steroid Hormones Steroid metabolite α-boldenone α-trenbolone β-boldenone β-trenbolone

This application note describes the extraction of a panel of 19 steroid hormones from human serum using EVOLUTE® EXPRESS ABN solid phase extraction plates prior to LC/MS analysis. The simple sample preparation procedure delivers clean extracts and analyte recoveries greater than 80% with RSDs lower than 10% for the majority of analytes. Linearity of >0.99 is achieved for all analytes in the range 5–5000 pg/mL. The application note includes details on automation of the extraction method using Biotage® Extrahera.

11-deoxycorticosterone 11-deoxycortisol 17-OH-Pregnenolone 17-alpha-hydroxyprogesterone 18-OH-Corticosterone 21-deoxycortisol 96-well plate Aldosterone Androstenedione Application Notes Clinical Corticosterone Cortisol Cortisone DHEA (dehydroepiandrosterone DHEAS Dehydroepiandrosterone (DHEA) Dihydrotestosterone (DHT) English Estradiol Estrogens Estrone Extrahera LC-MS/MS Pregnenolone Progesterone Serum Steroid Hormones Steroid metabolite

This application note describes the extraction of a panel of steroid hormones from human serum using ISOLUTE® SLE+ Supported Liquid Extraction plates prior to LC/MS-MS analysis. The simple sample preparation procedure delivers clean extracts and high, reproducible recoveries (>75%, RSD <10% ) for all analytes in human serum, with linearity >0.99 in the range 5–5000 pg/mL.

11-deoxycorticosterone 11-deoxycortisol 17-OH-Pregnenolone 17-alpha-hydroxyprogesterone 18-OH-Corticosterone 21-deoxycortisol 96-well plate Aldosterone Androstenedione Application Notes Clinical Corticosterone Cortisol Cortisone DHEA (dehydroepiandrosterone DHEAS Dehydroepiandrosterone (DHEA) Dihydrotestosterone (DHT) English Estradiol Estrogens Estrone LC-MS/MS Pregnenolone Progesterone Serum Steroid Hormones Steroid metabolite Testosterone

This application note describes the extraction of a panel of 19 steroid hormones from human urine using EVOLUTE® EXPRESS ABN solid phase extraction plates prior to LC-MS/MS analysis. The simple sample preparation procedure delivers clean extracts and analyte recoveries greater than 75% with RSDs lower than 5% for most analytes. Linearity of greater than 0.999 is achieved for all analytes in the range 1–1000 pg/mL. Manual and automated (using Biotage Extrahera) extraction protocols are detailed.

11-deoxycorticosterone 17-OH-Pregnenolone 17-alpha-estradiol 17-alpha-hydroxyprogesterone 17-beta-estradiol 18-OH-Corticosterone 21-deoxycortisol 96-well plate Aldosterone Androstenedione Application Notes Clinical Corticosterone Cortisol Cortisone DHEA (dehydroepiandrosterone DHEAS Dihydrotestosterone (DHT) English Estrone LC-MS/MS Pregnenolone Progesterone Steroid Hormones Steroid metabolite Testosterone Urine

This application note describes the extraction of a panel of 19 steroid hormones from human urine using ISOLUTE® SLE+ Supported Liquid Extraction plates prior to LC/MS-MS analysis. The simple sample preparation procedure delivers clean extracts and analyte recoveries greater than 90% with RSDs lower than 10% for all analytes. Linearity of greater than 0.999 is achieved for all analytes in the range 1-1000 pg/mL. Manual and automated (using Biotage Extrahera) extraction protocols are detailed.

11-deoxycorticosterone 11-deoxycortisol 17-OH-Pregnenolone 17-alpha-hydroxyprogesterone 18-OH-Corticosterone 21-deoxycortisol 96-well plate Aldosterone Androstenedione Application Notes Clinical Corticosterone Cortisol Cortisone DHEA (dehydroepiandrosterone DHEAS Dehydroepiandrosterone (DHEA) English Estradiol Estrone Extrahera LC-MS/MS Pregnenolone Progesterone Steroid Hormones Steroid metabolite Testosterone Urine

This poster demonstrates a rapid and reliable sample preparation method using Supported Liquid Extraction (ISOLUTE SLE+) to extract a suite of 30 hormone analytes from a hydrolyzed urine matrix. Single injection analysis by LC-MS/MS shows that matrix effects are eliminated by the ISOLUTE SLE+ protocol and that analyte recovery and sensitivity have excellent clinical utility. MSACL 2016

11-deoxycortisol 11-keto-androsterone 11-oxoetiocholanolone 11-β-hydroxyandrosterone 11β-hydroxyetiocholanolone 2-OH-estrone 2-methoxyestrone 3α-androstanediol 3β-androstanediol 4-OH-estrone 4-methoxyestrone 96-well plate Allo-THB Androsterone Clinical Corticosterone DHEA (dehydroepiandrosterone Dehydroepiandrosterone (DHEA) English Etiocholanolone Extrahera Hydrolysed urine Posters Steroid Hormones Steroid metabolite THB THF Testosterone α-THF