Development and Validation of a LC-MS/MS Assay for Quantification of Parathyroid Hormone (PTH 1-34) in human Plasma

Al Riyami, Sulaiman, Tang, Jonathan C Y ORCID:, Dutton, John, Washbourne, Christopher, Galitzer, Hillel and Fraser, William (2017) Development and Validation of a LC-MS/MS Assay for Quantification of Parathyroid Hormone (PTH 1-34) in human Plasma. In: ASBMR 2017 Annual Meeting, 2017-09-08 - 2017-09-11, Colorado Convention Center, ASBMR Discovery Hall.

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Background: Teriparatide [recombinant human PTH (1-34)] is an osteoanabolic agent for treatment of osteoporosis. The effect on bone decreases the risk of vertebral and non-vertebral fractures and increases bone mineral density (BMD) in post-menopausal women with osteoporosis. Measurement of PTH (1-34) is valuable in assessing treatment response and concordance with therapy.Aim: To develop and validate a method for quantification of PTH (1-34) using Liquid chromatography tandem mass spectrometry (LC-MS/MS) and to perform comparison with a commercial immunoassay.  Method: Sample extraction was developed using a Waters (Milford, MA, USA) Oasis® HLB µElution solid phase extraction. Quantification m/z transition 589>656 was used on Waters/Micromass® Quattro Ultima™ Pt mass spectrometer to measure PTH (1-34) in human plasma using rat PTH (1-34) as internal standard. Validation criteria were carried out against industry standards. PTH (1-34) results obtained from human subjects given Teriparatide (Fortsteo, Eli Lilly, IN, USA) (n=390) were compared against results obtained from an immunoassay (IDS; Boldon Tyne and Wear. UK).  Results and Discussion: LC-MS/MS produced a linear calibration curve from 10 to 2000 pg/mL (r2 >0.990). The LLoQ and LLoD for PTH (1-34) were 10 pg/mL and 2.1 pg/mL respectively. The inter- /intra-assay precision (CV%) of the method were <9.8 and <7.8% and accuracy of >98.3% for four QCs (20, 100, 200, and 800 pg/mL). The mean recovery of PTH (1-34) was 107.2%. Method comparison between the LC-MS/MS and immunoassay using human EDTA plasma samples showed a high correlation (r2 = 0.950). A concentration-dependent, negative bias of 35.5% was observed across the range of 0 – 800 pg/mL. The immunoassay showed a 7% cross reactivity to human PTH (1-84) and 44% to rat PTH (1-34), no interference was observed in the LC-MS/MS method. Matrix effect and cross reactivity to human PTH (1-84) in the immunoassay were the likely contributing factors to the bias between the methods. The oxidised form of PTH (1-34) does not interfere with our LC-MS/MS method.  Conclusion: Our LC-MS/MS method demonstrated linearity over the calibration range, good precision and accuracy, excellent analyte recovery, and negligible matrix effects. The method was successfully used for measurements of PTH (1-34) in rat and human plasma.

Item Type: Conference or Workshop Item (Poster)
Faculty \ School: Faculty of Medicine and Health Sciences > Norwich Medical School
UEA Research Groups: Faculty of Medicine and Health Sciences > Research Groups > Musculoskeletal Medicine
Faculty of Medicine and Health Sciences > Research Centres > Metabolic Health
Depositing User: Pure Connector
Date Deposited: 16 Nov 2017 06:09
Last Modified: 19 Oct 2023 04:00


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