LC-MS Method for measuring hormones in human serum

A Sensitive Liquid Chromatography–Tandem Mass Spectrometry Method for the Measurement of Nestorone in Human Serum

-Summary of MSACL 2017 Poster Presentation by: Feng Bai, Vignesh Iyer, Ronald Swerdloff, and Christina Wang; Los Angeles Biomedical Research Institute at Harbor-UCLA Medical Center

HybridSPE® Extraction

Nestorone® (NES) is a synthetic progestin being developed by Population Council and NICHD for female and male contraception.
NES does not bind to the androgen or the estrogen receptor and may have less adverse effects than androgenic progestin commonly used for female contraception.
NES was previously measured in serum by radioimmunoassay that has non specific interference leading to detectable NES levels in men when no NES was administered.
This objective of this study was to develop and validate a highly sensitive LC MS/MS method for the detection of serum Nestorone levels for research and clinical purposes in men.

Protein Precipitation	Serum Sample + I.S. + ACN with 1% Formic Acid Vortex & Centrifuge (for increased efficiency use Tip-on-Tip) Transfer supernatant
Condition	Aspirate and dispense HybridSPE- XTR Tips with 1000 μL acetonitrile
Extraction	Aspirate and dispense the supernatant 2x with HybridSPE- XTR Tips to bind interferents to sorbent Transfer remaining clean sample to a new glass vial
Prep for injection	Dry down the extracted samples Reconstitute with 40% MeOH in 0.1% formic acid H₂O for the analysis

Protein Precipitation

Serum Sample + I.S. + ACN with 1% Formic Acid
Vortex & Centrifuge (for increased efficiency use Tip-on-Tip)
Transfer supernatant

Condition

Aspirate and dispense HybridSPE- XTR Tips with 1000 μL acetonitrile

Extraction

Aspirate and dispense the supernatant 2x with HybridSPE- XTR Tips to bind interferents to sorbent

Transfer remaining clean sample to a new glass vial

Prep for injection

Dry down the extracted samples

Reconstitute with 40% MeOH in 0.1% formic acid H₂O for the analysis

You can automate this method with Tip-on-Tip for protein precipitation and a HybridSPE XTR tip format for an automated liquid handler.

sample preparation for blood/ serum samples prior to LC-MS/MS analysis

INTip Solutions

Tip-on-Tip

Combines a top wide bore conductive tip and a bottom tip for filtration. Filtration tips are available on Hamilton and Integra platforms. Tip-on-Tip technology was designed to automate and simplify complex sample preparation.

Dispersive Pipette XTRaction

XTR tips can contain loose HybridSPE sorbent material in 30 mg or 50 mg depending on the tip format. Tips are available for Tecan, Hamilton, Integra and Universal style for multi-channel pipettors.