Vasanta Putluri (Presenter)
Baylor College of Medicine
Bio: I joined Baylor College of Medicine as an Lab Manager in 2011. I have published 5 peer reviewed papers of which 4 papers are in the area of Cancer Metabolomics.
Authorship: Vasanta Putluri, Andre Szczesniewski, Vadiraj Bhat, Feng Jin, Peter J Cook, SriRamya Denepudi, Dolores J Lamb, Arun Sreekumar and Putluri Nagireddy
Baylor College of Medicine, Houston, TX, Agilent Technology
| Short Abstract Androgens (such as testosterone) are natural hormones. They are important in sexual development in both men and women. In women, androgens are produced in small amounts by the ovaries and the adrenal glands. Similar to higher blood estrogen levels, higher amounts of androgens in the blood may be linked to an increased risk of breast cancer in women. An ultra sensitive quantitative analytical method is required for total testosterone in human serum to be able to measure the low levels found in women. Therefore, a method was developed on an Agilent 6495 Ion Funnel Mass Spectrometer to quantify and characterize testosterone underivatized in serum and to ascertain which method may be the most appropriate for laboratory use. |
Long Abstract
Androgens (such as testosterone) are natural hormones. They are important in sexual development in both men and women. In women, androgens are produced in small amounts by the ovaries and the adrenal glands. Similar to higher blood estrogen levels, higher amounts of androgens in the blood may be linked to an increased risk of breast cancer in women. Studies show higher blood levels of testosterone may increase the risk of breast cancer in postmenopausal women. We therefore developed a sensitive and specific stable-isotope dilution liquid chromatography-tandem mass spectrometric (LC-MS/MS) method for serum testosterone at the concentrations encountered in human serum. Testosterone was extracted with ether-ethyl acetate from 100 uL of serum. Instrumental analysis was performed on an Agilent 6495 QQQ mass spectrometer in the multiple-reaction monitoring (MRM) mode after separation on a Agilent poroshell 120SB C18 reversed-phase column. The MRM transitions (m/z) were 289/97 for testosterone and 292/100 for [13C]3 testosterone. The calibration curves exhibited consistent linearity and repeatability in the range 200–400 ng/DL. Interassay CVs were <5 % at mean concentrations of testosterone of 200–400 ng/DL. The limit of detection was 2.5 pg/mL (signal-to-noise ratio 5) and the overall method recovery of testosterone was 90 %. Correlation (r) with our in-house extraction RIA was 0.98.. Sensitivity and specificity of the LC-MS/ MS method correlate with ELISA and make it suitable for use as a high-throughput clinical research method
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