An in Vitro Comparison of Blood Collection Tubes: Identifying Tubes Which Minimize Interference of Testosterone Measurement By Testosterone Ester

Program: Abstracts - Orals, Poster Previews, and Posters
Session: SUN 176-202-Male Reproductive Endocrinology and Male Reproductive Tract (posters)
Bench to Bedside
Sunday, April 3, 2016: 1:15 PM-3:15 PM
Exhibit/Poster Hall (BCEC)

Poster Board SUN 188
Jonas Ceponis*1, Andrew Leung1, Ronald S. Swerdloff1, Robert E. Dudley2, Theodore M. Danoff2 and Christina Wang1
1LABioMed at Harbor-UCLA Medical Center, Torrance, CA, 2Clarus Therapeutics, Inc, Northbrook, IL
Background: Non-specific esterases can hydrolyze steroid esters after blood collection leading to inaccurate steroid concentration results. Objective: Our goal was to determine which blood collection tube (i.e., which tube additives) would least interfere with testosterone (T) measurements in the presence of T Undecanoate (TU). Methods: In each of 4 experiments, 50 ml of venous blood was collected from healthy volunteers and 2 ml were immediately transferred into duplicate blood collection tubes including plain tube (red top), NaF (sodium fluoride 30 mg), P-800 tube (with esterase/protease inhibitors), EDTA, NaF 3mg+EDTA, or NaF 10mg+Oxalate (all manufactured by BD Diagnostics, Franklin Lakes, NJ). Known amounts of TU (125 to 1000 ng/ml in methanol) were added to each tube either before or after blood was collected. Prior to centrifugation the tubes were stored for 30 or 60 minutes at 4ºC or at room temperature (RT). Serum or plasma T levels were measured by LC-MS/MS using the Shimadzu high-performance LC 20 series system (Columbia, MD) with an Applied Biosystems API 5500 with ESI source (Foster City, CA) as previously described (Shiraishi, et al., 2008). Results: T levels in plain tube with no TU added were used as the reference value for all experiments. All tubes exhibited a TU dose related increase in T levels presumably by non-specific esterases in blood. Blood collected in plain, EDTA or P800 tubes after addition of TU 600 ng/ml led to an overestimation of +28% and +259% for plain, +28% and +318% for EDTA, and +22% and +227% for P800 tubes for 30 min at 4ºC and at 60 min at RT, respectively). Tubes containing NaF led to a lesser increase in serum T levels because presence of NaF resulted in underestimation of T levels even when no TU was present (up to -23%, - 10%, -3%, for NaF 30 mg, NaF10 mg +Oxalate, and NaF 3mg +EDTA at 4ºC, respectively). When TU 600 ng/ml was added to these tubes with NaF, serum T was changed by -4%, +4%, +14% for NaF 30 mg, NaF 10mg +Oxalate, and NaF 3mg +EDTA at 4ºC for 30min, respectively). Conclusion: The least interference by TU on T measurement was observed when tubes were stored for 30 minutes at 4ºC before centrifugation. NaF reduced hydrolysis of TU but inherently decreased T measured in blood. For tubes containing NaF, the extent of underestimation as well as inhibition of hydrolysis depends on amount of NaF in the tube.

(1) Shiraishi S, Lee P W, Leung A, Goh V H, Swerdloff R S & Wang C. (2008) Simultaneous measurement of serum testosterone and dihydrotestosterone by liquid chromatography-tandem mass spectrometry. Clin. Chem. 54, 1855-1863.

Disclosure: RED: Employee, Clarus, Employee, Clarus, Employee, Clarus. TMD: Employee, Clarus, Employee, Clarus, Former employee, GlaxoSmithKline, Former employee, Endo Pharmaceuticals. CW: Principal Investigator, Clarus, Principal Investigator, Lipocine, Principal Investigator, Besins HealthCare, Principal Investigator, Prolor, Advisory Group Member, Lipocine, Advisory Group Member, TesoRX. Nothing to Disclose: JC, AL, RSS

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