Testosterone

No overview yet.

Method comparison

Beckman Access/DxI 9000

format competitive use quantitative AMR Approximately 0.4 - 16.0 ng/mL (1.39 - 55.5 nmol/L) LoD 0.3 ng/mL (0.9 nmol/L) maximum observed; design criteria ≤ 0.4 ng/mL (≤ 1.39 nmol/L) LoQ 0.3 ng/mL (0.9 nmol/L) maximum observed at ≤ 20% within-lab CV; design criteria ≤ 0.4 ng/mL (≤ 1.39 nmol/L) LoB 0.2 ng/mL (0.7 nmol/L) maximum observed; design criteria ≤ 0.2 ng/mL (≤ 0.7 nmol/L)
[Paramagnetic bead]-Goat anti-mouse IgG (polyclonal) [conjugate bridge] :: [Capture mAb, mouse anti-testosterone, monoclonal] :: Testosterone [analyte, free hapten released from SHBG/albumin, monovalent]
[Paramagnetic bead]-Goat anti-mouse IgG (polyclonal) [conjugate bridge] :: [Capture mAb, mouse anti-testosterone, monoclonal] :: Testosterone-[Alkaline phosphatase label] [competitor analog occupying the same monovalent site as the analyte]
  • R1a — Particle/Conjugate reagent: Paramagnetic particles coated with goat anti-mouse IgG; testosterone alkaline phosphatase conjugate with bovine serum albumin (BSA), < 0.1% sodium azide, 0.1% ProClin 300
  • R1b — Sample Treatment Solution: Acetic acid < 5%, < 0.1% sodium azide
  • R1c — Anti-testosterone antibody reagent: Monoclonal anti-testosterone (mouse), protein (BSA, mouse, goat), < 0.1% sodium azide, 0.1% ProClin 300
  • Beads:
    • Base material: Paramagnetic particles
    • Coating: Goat anti-mouse IgG (polyclonal)
  • Capture reagent:
    • Host: mouse
    • Clonality: monoclonal
  • Conjugate/bridge reagent:
    • Host: goat
    • Clonality: polyclonal
    • Molecule type: IgG
  • Competitor analog:
    • Molecule type: Non-Ab: testosterone hapten conjugated to an enzyme label, not an antibody
    • Conjugate/label: Alkaline phosphatase
  • Sample pretreatment:
    • Composition: Acetic acid < 5%, < 0.1% sodium azide
    • Purpose: Dissociates testosterone from its serum carrier proteins (SHBG, albumin) before the competitive binding reaction, releasing total (bound + free) testosterone for measurement
  • Wash buffer:
    • Purpose: Washes away unbound conjugate, unbound patient testosterone, and other unbound reagents after magnetic separation of the solid phase

Reaction procedure:

  1. Sample (20 µL) is added to the reaction vessel together with R1b (Sample Treatment Solution), R1c (mouse monoclonal anti-testosterone antibody), and R1a (paramagnetic particles coated with goat anti-mouse IgG, co-formulated with testosterone-alkaline phosphatase conjugate) — all four are combined and co-incubated in a single one-step addition, with no intervening wash before the binding competition, consistent with the assay's stated 'one-step, competitive' design.
  2. Within R1b, acetic acid (< 5%) dissociates testosterone from its serum carrier proteins (SHBG, albumin), releasing total testosterone so it is available to compete with the labeled conjugate.
  3. Released patient testosterone and the testosterone-alkaline phosphatase conjugate (from R1a) compete for a limited number of binding sites on the mouse monoclonal anti-testosterone antibody (R1c) — the antibody is present in limiting amount, which is what makes the reaction competitive rather than a two-site sandwich.
  4. Whichever species occupies the antibody's binding site (patient testosterone or the ALP-conjugate) is carried, via the antibody, onto the paramagnetic particles: the particles' goat anti-mouse IgG coating (from R1a) binds the mouse antibody itself regardless of what it is holding, bridging the whole complex to the solid phase.
  5. The vessel is placed in a magnetic field; solid-phase-bound material (particles plus bound antibody and whatever it captured) is retained while unbound conjugate, unbound patient testosterone, and other unbound reagents are washed away.
  6. Chemiluminescent substrate (Lumi-Phos PRO) is added; alkaline phosphatase on the fraction of conjugate that ended up bead-bound generates light, measured by a luminometer. Because bead-bound conjugate and patient testosterone compete for the same limited antibody sites, light output is inversely proportional to the patient's testosterone concentration.
  • What's measured: Measures total testosterone (protein-bound plus free) — the Sample Treatment Solution (R1b) chemically releases testosterone from SHBG and albumin before the competitive reaction, so results are not free testosterone. The cross-reactivity table shows modest reactivity with some closely related steroids (11-ketotestosterone 6.7%, 11β-hydroxytestosterone 4.1%, 5-alpha-DHT 2.0%, 19-nortestosterone 1.6%, mesterolone 1.5%), which could contribute a small positive bias if these are elevated (e.g., anabolic steroid use), while cross-reactivity with cortisol, estradiol, progesterone, DHEA, and other major steroids/steroid conjugates tested is negligible (≤ 0.7%).
  • Interference implications: Hemolysis, lipemia (triglycerides), icterus (bilirubin), and total protein across the ranges tested do not significantly affect results. The insert separately flags heterophile antibodies (HAMA, human anti-goat antibodies — particularly relevant since capture depends on a goat anti-mouse bridge), rheumatoid factor, endogenous alkaline phosphatase, fibrin, and ALP-binding proteins as potential (but not quantified or insert-specific) sources of erroneous results; no direction of bias is stated for these, so unexpected or clinically discordant results should prompt evaluation for interfering antibodies or ALP-related interferents.
  • Intended use: Reported as a quantitative total testosterone concentration (ng/mL, convertible to nmol/L by × 3.47); no reflex or confirmatory testing is indicated by this insert. Results below the LoQ (~0.4 ng/mL) are reported as 'less than,' and results above the top calibrator (16.0 ng/mL) as 'greater than' or via a 1:1 dilution with Calibrator S0. Interpret against the laboratory's own established reference interval and the patient's full clinical picture, per Endocrine Society guidance for evaluating androgen deficiency.
  • Known bias: Method comparison of Access 2 vs. DxI 9000 (CLSI EP09c, N=108, range 0.48-14 ng/mL, Weighted Deming regression) gave slope 0.95 (95% CI 0.93-0.98), intercept 0.028 (95% CI -0.015 to 0.071), R=0.99 — DxI 9000 reads modestly lower than Access 2 (~5% low slope) with negligible intercept bias.
Interferences & pre-/post-analytical notes

Cross-reactivity

  • Testosterone-glucuronide: 0.4% cross-reactivity (tested at 100 ng/mL)
  • Testosterone-sulfate: 0.3% cross-reactivity (tested at 100 ng/mL)
  • 5-alpha-DHT: 2.0% cross-reactivity (tested at 100 ng/mL)
  • Androstanediol: 0.4% cross-reactivity (tested at 100 ng/mL)
  • Androstenediol: 0.6% cross-reactivity (tested at 100 ng/mL)
  • Androstenedione: 0.7% cross-reactivity (tested at 100 ng/mL)
  • DHEA: 0.0% cross-reactivity (tested at 1,000 ng/mL)
  • DHEA-sulfate: 0.0% cross-reactivity (tested at 1,000 ng/mL)
  • Androsterone: 0.2% cross-reactivity (tested at 100 ng/mL)
  • Corticosterone: 0.0% cross-reactivity (tested at 1,000 ng/mL)
  • Cortisol: 0.0% cross-reactivity (tested at 1,000 ng/mL)
  • Estradiol: 0.0% cross-reactivity (tested at 100 ng/mL)
  • Estradiol-sulfate: 0.0% cross-reactivity (tested at 100 ng/mL)
  • Estriol: 0.2% cross-reactivity (tested at 100 ng/mL)
  • Estrone: 0.4% cross-reactivity (tested at 100 ng/mL)
  • Estrone-glucuronide: 0.0% cross-reactivity (tested at 100 ng/mL)
  • Estrone-sulfate: 0.0% cross-reactivity (tested at 100 ng/mL)
  • Progesterone: 0.4% cross-reactivity (tested at 100 ng/mL)
  • 11-Deoxycortisol: 0.0% cross-reactivity (tested at 1,000 ng/mL)
  • 17-alpha-Hydroxyprogesterone: 0.1% cross-reactivity (tested at 100 ng/mL)
  • 19-Hydroxytestosterone: 0.5% cross-reactivity (tested at 100 ng/mL)
  • 2-Hydroxyestradiol: 0.0% cross-reactivity (tested at 100 ng/mL)
  • Ethinylestradiol: 0.3% cross-reactivity (tested at 100 ng/mL)
  • Mestranol: 0.0% cross-reactivity (tested at 100 ng/mL)
  • Norethindrone: 0.05% cross-reactivity (tested at 100 ng/mL)
  • Norgestrel: 0.3% cross-reactivity (tested at 100 ng/mL)
  • Danazol: 0.3% cross-reactivity (tested at 100 ng/mL)
  • Mesterolone: 1.5% cross-reactivity (tested at 100 ng/mL)
  • Dexamethasone: 0.0% cross-reactivity (tested at 1,000 ng/mL)
  • 19-Nortestosterone: 1.6% cross-reactivity (tested at 100 ng/mL)
  • Ethinyltestosterone: 0.07% cross-reactivity (tested at 100 ng/mL)
  • 19-Norethisterone Acetate: 0.02% cross-reactivity (tested at 100 ng/mL)
  • 11B-Hydroxytestosterone: 4.1% cross-reactivity (tested at 100 ng/mL)
  • 11-Ketotestosterone: 6.7% cross-reactivity (tested at 100 ng/mL)
  • 17-alpha-Methyltestosterone: 0.2% cross-reactivity (tested at 100 ng/mL)

Interferences

Suspected / theoretical
  • Heterophile antibodies (HAMA, human anti-goat antibodies): May cause erroneous results; risk raised specifically because the assay uses animal-derived antibodies including a goat anti-mouse solid-phase reagent — Kricka L. Interferences in immunoassays - still a threat. Clin Chem 2000; 46: 1037-1038; Bjerner J, et al. Immunometric assay interference: incidence and prevention. Clin Chem 2002; 48: 613-621.
  • Rheumatoid factor: Documented in literature as a potential cause of erroneous immunoassay results
  • Endogenous alkaline phosphatase: Documented in literature as a potential cause of erroneous results in assays using an alkaline phosphatase label
  • Fibrin: Documented in literature as a potential cause of erroneous immunoassay results
  • Proteins capable of binding to alkaline phosphatase: May cause erroneous results in this ALP-labeled assay — Lingwood D, Ballantyne JS. Alkaline phosphatase-immunoglobulin conjugate binds to lipids in vitro, independent of antibody selectivity. Journal of Immunological Methods 2006; 311: 174-177.
Validated -- no significant interference
  • Bilirubin: Does not significantly affect the concentration of total testosterone assayed (Up to 10 mg/dL (171 µmol/L))
  • Hemoglobin (hemolysis): Does not significantly affect the concentration of total testosterone assayed (Up to 1,000 mg/dL (10 g/L))
  • Triglycerides (Triolein): Does not significantly affect the concentration of total testosterone assayed (Equivalent of up to 1,800 mg/dL (20.32 mmol/L))
  • Total protein (human serum albumin): Does not significantly affect the concentration of total testosterone assayed (5.5–8.5 g/dL)

Pre-analytical

  • Serum and plasma (heparin) are the recommended sample types; EDTA plasma is not recommended and has been shown to give erroneous results — do not use EDTA plasma.
  • Serum and plasma values should not be used interchangeably; use the reference interval specific to the sample type collected.
  • Store samples tightly stoppered at room temperature (15-30°C) for no longer than 8 hours; refrigerate at 2-8°C if not completed within 8 hours; freeze at -20°C or colder if not completed within 48 hours or for shipment.
  • Samples may be thawed and refrozen up to two times.

Post-analytical

  • Results below the lower limit of the measuring interval are reported as less than that value; results above the highest calibrator (16.0 ng/mL) are reported as greater than that value, or the sample may be diluted 1:1 with Access Testosterone Calibrator S0 and re-tested, with the system adjusting the result for dilution.
  • Testosterone results from different manufacturers' assays will vary; per Endocrine Society guidance, evaluate androgen deficiency against the lower limit of the reference interval established by the testing laboratory, in light of the patient's full clinical presentation.
Source: package insert (PDF, Cat #33560, rev C85437 F (Revision F, January 2026)) · Extracted 2026-08-26 · Approved 2026-09-10

Roche cobas e801

format competitive use quantitative AMR 2.50-1500 ng/dL or 0.087-52.0 nmol/L LoD 2.50 ng/dL or 0.087 nmol/L LoQ 12.0 ng/dL or 0.416 nmol/L LoB 1.50 ng/dL or 0.052 nmol/L
[Streptavidin-coated microparticle] :: Biotin-[Capture mAb] :: Testosterone-peptide~Ru(bpy)3 [competitor analog, signal-generating, bound instead of analyte]
[Streptavidin-coated microparticle] :: Biotin-[Capture mAb] :: Testosterone [analyte, monomeric, released from SHBG, bound instead of labeled analog]
  • M — Streptavidin-coated microparticles (solid phase): Streptavidin-coated microparticles 0.72 mg/mL; preservative
  • R1 — Biotinylated anti-testosterone antibody with releasing reagent: Biotinylated monoclonal anti-testosterone antibody (sheep) 40 ng/mL; releasing reagent 2-bromoestradiol; MES buffer 50 mmol/L, pH 6.0; preservative
  • R2 — Ruthenium-labeled testosterone derivative (competitor): Testosterone derivative, labeled with ruthenium complex 1.5 ng/mL; MES buffer 50 mmol/L, pH 6.0; preservative
  • Beads:
    • Coating: streptavidin
  • Capture reagent:
    • Host: sheep
    • Clonality: monoclonal
    • Conjugate/label: biotin
  • Competitor analog:
    • Molecule type: Non-Ab small-molecule analyte analog -- testosterone derivative conjugated to a peptide linker and a ruthenium label, not a binder protein
    • Conjugate/label: Ruthenium complex (Ru(bpy)3, tris(2,2'-bipyridyl)ruthenium(II))
  • Sample releasing reagent (component of R1):
    • Composition: 2-bromoestradiol, in MES buffer 50 mmol/L pH 6.0 (within R1)
    • Purpose: Displaces endogenous testosterone bound to serum carrier proteins (SHBG), releasing it so it is free to bind the antibody -- enables measurement of total rather than only free testosterone
  • Measuring-cell wash solution (system reagent, ProCell II M):
    • Purpose: Removes unbound sample and reagent components from the magnetically captured microparticles before electrochemiluminescent read, so only bead-bound complex contributes to signal

Reaction procedure:

  1. First incubation: 12 uL of sample is combined with R1 (biotinylated anti-testosterone monoclonal antibody, which also carries the releasing reagent 2-bromoestradiol). The 2-bromoestradiol displaces testosterone from serum carrier proteins (SHBG), freeing endogenous testosterone to occupy binding sites on the biotinylated antibody in proportion to its concentration -- this equilibration is allowed to proceed before any labeled reagent is introduced.
  2. Second incubation: M (streptavidin-coated microparticles) and R2 (Ru(bpy)3-labeled testosterone derivative) are added together. R2 competes for whatever antibody binding sites were left unoccupied by endogenous testosterone in step 1; the resulting antibody complex (bound to either endogenous testosterone or R2) becomes localized to the microparticles via the biotin tag on the antibody docking onto the bead's streptavidin coating.
  3. The reaction mixture is aspirated into the measuring cell, where the microparticles are magnetically captured on the electrode surface; unbound sample/reagent components are washed away with ProCell II M, removing any R2 that did not end up antibody/bead-bound.
  4. A voltage is applied to the electrode, inducing electrochemiluminescent emission from the Ru(bpy)3 label on bead-bound complexes, measured by a photomultiplier. Because R2 and endogenous testosterone compete for the same limited antibody sites, signal intensity is inversely proportional to sample testosterone concentration.
  5. The analyzer converts the ECL signal to a testosterone concentration via an instrument-specific 2-point calibration applied to a reagent-lot master/leading calibration curve delivered through cobas link.
  • What's measured: Measures total testosterone (protein-bound plus free), since the releasing reagent 2-bromoestradiol in R1 dissociates testosterone from SHBG before the antibody-binding step. The assay does not distinguish endogenous testosterone from exogenous/metabolized testosterone (e.g., testosterone undecanoate therapy). Some cross-reactivity exists with structurally related steroids, most notably 11-beta-hydroxy-testosterone (up to ~20.6%) and 11-keto-testosterone (up to ~4.87%), with lower-level cross-reactivity to androstenedione, ethisterone, and several other steroids; cortisol, cortisone, dexamethasone, estrone, prednisone, and prednisolone show no detectable cross-reactivity even at high concentrations.
  • Interference implications: Bilirubin, hemoglobin, lipemia (Intralipid), biotin, and rheumatoid factor all show no impact on results at the tested thresholds (biotin validated clean up to 1200 ng/mL, above the ~1160 ng/mL seen after very high single-dose biotin supplementation). Phenylbutazone at therapeutic dosage causes falsely increased testosterone results. Nandrolone shows a strong cross-reaction and samples from patients on Nandrolone therapy should not be tested. Rare heterophile/anti-reagent antibodies (to the analyte-specific antibody, streptavidin, or ruthenium) may cause interference at extremely high titers, and isolated cases of falsely elevated results have been seen in female ESRD patients -- both should prompt confirmation by an orthogonal method if results seem implausible.
  • Intended use: Result is reported as a continuous quantitative testosterone concentration (ng/dL, ng/mL, or nmol/L) rather than a qualitative or S/CO call. Because this is a single-antibody competitive format rather than a two-antibody sandwich, and because the assay does not distinguish endogenous from exogenous/metabolized testosterone, the insert recommends confirming implausibly elevated results (particularly in women) with an extraction method or a validated LC-MS/MS assay.
  • Known bias: Comparison of the current Elecsys Testosterone II reagent (08946370190) against the prior version (07027915190) on the cobas e801, n=169, gave Passing/Bablok y=1.02x-2.71 (tau=0.980) and linear regression y=1.02x-6.04 (r=0.999) over a sample range of 4.61-1400 ng/dL, indicating close agreement between reagent lots/versions.
Interferences & pre-/post-analytical notes

Cross-reactivity

  • Androstenedione: <= 3.15% cross-reactivity (tested at 100 ng/mL)
  • Cortisol: not detectable cross-reactivity (tested at 5000 ng/mL)
  • Cortisone: not detectable cross-reactivity (tested at 5000 ng/mL)
  • Danazol: <= 0.504% cross-reactivity (tested at 1000 ng/mL)
  • Dexamethasone: not detectable cross-reactivity (tested at 5000 ng/mL)
  • DHEA: <= 0.014% cross-reactivity (tested at 5000 ng/mL)
  • DHEA-S: <= 0.003% cross-reactivity (tested at 50000 ng/mL)
  • D-5-Androstene-3beta,17beta-diol: <= 0.289% cross-reactivity (tested at 1000 ng/mL)
  • Estradiol: <= 0.211% cross-reactivity (tested at 5000 ng/mL)
  • Estrone: not detectable cross-reactivity (tested at 5000 ng/mL)
  • Ethisterone: <= 3.57% cross-reactivity (tested at 300 ng/mL)
  • Norgestrel: <= 0.539% cross-reactivity (tested at 1000 ng/mL)
  • Testosterone propionate: <= 0.718% cross-reactivity (tested at 100 ng/mL)
  • 5-alpha-Androstane-3beta,17beta-diol: <= 2.15% cross-reactivity (tested at 500 ng/mL)
  • 5-alpha-Dihydro-testosterone: <= 1.30% cross-reactivity (tested at 500 ng/mL)
  • 11-beta-Hydroxy-testosterone: <= 20.6% cross-reactivity (tested at 50 ng/mL)
  • 11-Keto-testosterone: <= 4.87% cross-reactivity (tested at 200 ng/mL)
  • 19-Norethisterone: <= 5.51% cross-reactivity (tested at 40 ng/mL)
  • Prednisone: not detectable cross-reactivity (tested at 5000 ng/mL)
  • Prednisolone: not detectable cross-reactivity (tested at 5000 ng/mL)
  • Progesterone: <= 0.009% cross-reactivity (tested at 5000 ng/mL)

Interferences

Known interferences
  • Phenylbutazone: Testosterone values falsely increased at therapeutic dosage levels (107 mg/L)
  • Nandrolone: Strong interaction/cross-reaction; do not use samples from patients under Nandrolone treatment (11.4 mg/L)
Suspected / theoretical
  • Testosterone Undecanoate (and other exogenous testosterone therapy): Metabolized to testosterone in vivo; assay does not differentiate endogenous from exogenous/metabolized testosterone, so results reflect total testosterone including supplementation (32 mg/L tested; recovery within +/-10% criterion)
  • Antibodies to analyte-specific antibodies, streptavidin, or ruthenium (heterophile/anti-reagent antibodies): May rarely interfere at extremely high titers; minimized by test design
  • End-stage renal disease (ESRD) patient status, female patients: Isolated cases of falsely/implausibly elevated testosterone reported; confirm with an extraction method or validated LC-MS/MS assay — Rosner W et al., J Clin Endocrinol Metab 2007;92(2):404-413
Validated -- no significant interference
  • Bilirubin: No impact on results (<= 513 umol/L or <= 30 mg/dL)
  • Hemoglobin: No impact on results (<= 0.373 mmol/L or <= 600 mg/dL)
  • Intralipid (lipemia): No impact on results (<= 800 mg/dL)
  • Biotin: No biotin interference observed; patients on high-dose biotin supplementation (e.g. up to 300 mg single dose) can reach serum levels up to ~1160 ng/mL, close to but still under the tested ceiling (<= 4912 nmol/L or <= 1200 ng/mL) — Grimsey P et al., Int J Pharmacokinet 2017;2(4):247-256; Piketty ML et al., Clin Chem Lab Med 2017;55(6):817-825
  • Rheumatoid factors: No impact on results (<= 1000 IU/mL)

Pre-analytical

  • Acceptable specimens: serum (standard or separator-gel tubes), and Li-heparin, K2-EDTA, or K3-EDTA plasma.
  • Sample stability: 14 days at 2-8C, 5 days at 20-25C, 6 months at -20C (+/-5C); freeze only once.
  • Do not use heat-inactivated samples.
  • Do not use samples or controls stabilized with azide.
  • Centrifuge samples containing precipitates before assay.
  • Analyze samples and calibrators within 2 hours of reaching 20-25C due to evaporation effects.
  • Do not use samples from patients under Nandrolone treatment (strong assay interference).

Post-analytical

  • Values below the Limit of Detection are reported as < 2.50 ng/dL (< 0.087 nmol/L); values above the measuring range are reported as > 1500 ng/dL (> 52.0 nmol/L).
  • Results < 12.0 ng/dL should be annotated as not reliable, since intermediate precision CV exceeds 20% below the Limit of Quantitation.
  • Implausibly elevated testosterone values in women should be verified by an extraction method or a validated LC-MS/MS tandem method.
Source: package insert (PDF, Cat #09745963190, 09745963501, rev 2026-04, V 1.0) · Extracted 2026-08-26 · Approved 2026-09-10

Siemens Atellica IM

format competitive use quantitative AMR 7.00–1500.00 ng/dL (0.24–52.05 nmol/L); reportable range extendable to 3000.00 ng/dL (104.10 nmol/L) via 1:2 automated dilution with Multi-Diluent 3 LoD Design goal ≤ 5.00 ng/dL (0.17 nmol/L); representative value 4.43 ng/dL (0.15 nmol/L), determined per CLSI EP17-A2 using 320 determinations (256 blank, 64 low-level replicates) LoQ Design goal ≤ 7.00 ng/dL (0.24 nmol/L); representative value 4.43 ng/dL (0.15 nmol/L), defined as the lowest concentration where within-laboratory CV ≤ 20% LoB Design goal ≤ 2.50 ng/dL (0.087 nmol/L); representative value 1.81 ng/dL (0.06 nmol/L)
[Latex particle]-Streptavidin :: Biotin-[Capture mAb] :: Acridinium ester-[Competitor analog, testosterone hapten]
[Latex particle]-Streptavidin :: Biotin-[Capture mAb] :: Testosterone [analyte, monomeric, released from binding proteins]
  • Releasing Agent — Steroid releasing agent + biotinylated anti-testosterone antibody: Steroid releasing agent (0.4 µg/mL); biotinylated sheep monoclonal anti-testosterone antibody (27 µg/L) in buffered saline; preservatives
  • Lite Reagent — Acridinium ester-labeled testosterone hapten: Acridinium ester-labeled hapten (36 µg/mL) in buffered saline; preservatives
  • Solid Phase — Streptavidin-coated latex particles: Streptavidin-coated latex particles (0.33 g/L) in buffered saline; preservatives
  • Atellica IM Wash — System wash buffer:
  • Acid — Chemiluminescent trigger reagent (acid):
  • Base — Chemiluminescent trigger reagent (base):
  • Multi-Diluent 3 — Automated sample dilution diluent: Human plasma; sodium azide (0.1%)
  • APW1 — Probe wash: 0.4 N sodium hydroxide
  • Beads:
    • Base material: latex particle
    • Coating: streptavidin
  • Capture reagent:
    • Host: sheep
    • Clonality: monoclonal
    • Conjugate/label: biotin
  • Competitor analog:
    • Molecule type: Non-Ab, testosterone hapten analog (small-molecule steroid conjugate)
    • Conjugate/label: acridinium ester
  • Sample pretreatment/release reagent (within Releasing Agent):
    • Composition: Steroid releasing agent 0.4 µg/mL in buffered saline
    • Purpose: Dissociates testosterone from endogenous binding proteins (SHBG/TeBG, CBG, albumin) so total (bound + unbound) testosterone becomes available to bind the anti-testosterone antibody
  • Wash buffer (post competitive-binding incubation, step 3):
    • Purpose: Removes unbound Lite Reagent (free acridinium-labeled hapten) and unbound sample matrix before the chemiluminescent trigger, so only the bead-bound antibody-label fraction contributes signal
  • Sample dilution diluent (automated dilution for samples > 1500 ng/dL):
    • Composition: Human plasma; sodium azide (0.1%)
  • Probe wash (system fluid, not part of the assay reaction chemistry):
    • Composition: 0.4 N sodium hydroxide

Reaction procedure:

  1. Dispense 20 µL sample + 90 µL Releasing Agent into the cuvette; co-incubate 9 min at 37°C. The steroid releasing agent dissociates testosterone from endogenous SHBG/TeBG, CBG, and albumin, freeing total testosterone; simultaneously the biotinylated sheep monoclonal anti-testosterone antibody in Releasing Agent begins binding the now-free patient testosterone, establishing the competitive equilibrium before the labeled competitor is introduced.
  2. Dispense 50 µL Lite Reagent + 150 µL Solid Phase together; co-incubate 3 min at 37°C. Acridinium ester-labeled testosterone hapten in Lite Reagent now competes with any patient testosterone for antibody sites not already occupied; whichever species (patient testosterone or labeled hapten) is antibody-bound gets pulled onto the Solid Phase's streptavidin coating via the antibody's biotin tag, localizing only antibody-bound complexes to the bead. Patient testosterone and labeled hapten therefore compete for a fixed pool of antibody, so the amount of labeled hapten ending up bead-bound is inversely related to patient testosterone concentration.
  3. Separate, aspirate, and wash the cuvette with Atellica IM Wash, removing unbound (free) Lite Reagent and other unbound sample/reagent components -- what remains bead-bound is only the antibody-label complex fraction.
  4. Dispense 300 µL each of Acid and Base to initiate the chemiluminescent flash reaction from the acridinium ester label remaining on the bead.
  5. Report RLU and convert to testosterone concentration via the master curve; RLU is inversely proportional to patient testosterone.
  • What's measured: Measures total testosterone (bound + unbound) -- the Releasing Agent's steroid releasing agent actively strips testosterone off SHBG/TeBG, CBG, and albumin before the competitive binding step, so the result reflects total circulating hormone, not the free/bioavailable fraction (that requires calculating FAI against SHBG separately). Specificity for testosterone itself is high, but the anti-testosterone antibody meaningfully cross-reacts with closely related steroid analogues -- notably 11β-hydroxytestosterone (~15%) and, to a lesser extent, 11-keto-testosterone and testosterone propionate -- and cannot be used at all in patients on nandrolone decanoate.
  • Interference implications: Biotin is the dominant interference risk: results stay within 10% up to 30 ng/mL biotin, but bias climbs steeply above that (>80% bias by 250-500 ng/mL) and drives falsely elevated results -- clinically relevant given that high-dose biotin supplement users can reach 100s of ng/mL. Bilirubin behaves asymmetrically and is described as causing 'erroneous' (not simply elevated) results: conjugated bilirubin biases results downward (-14.4% at 20 mg/dL) while unconjugated bilirubin biases upward (+20.1% at 30 mg/dL), so icteric samples above the stated thresholds should be flagged rather than assumed to skew in one direction. Heterophile/HAMA antibodies are a suspected but not fully excluded risk -- the assay is designed to minimize this interference but can still yield falsely elevated or falsely depressed results in individual patients. Hemolysis, lipemia, cholesterol, and rheumatoid factor were all validated as non-interfering up to the tested levels.
  • Intended use: Reports a quantitative total testosterone concentration for evaluating androgen-related disorders (e.g., male hypogonadism, female hirsutism/virilization, precocious/delayed puberty); the insert states results should always be interpreted alongside the patient's medical history and clinical presentation, with additional/confirmatory testing suggested when results are inconsistent with clinical evidence (particularly relevant given the assay's cross-reactivity with structurally related steroids). The total testosterone value can be paired with an SHBG result to derive the Free Androgen Index (FAI) as an estimate of physiologically active testosterone.
  • Known bias: Excellent agreement with the predicate ADVIA Centaur TSTII assay (Deming regression: y = 0.97x + 2.51 ng/dL, r = 1.00, n=111). Also correlates well with an ID-LC-MS/MS reference method (CDC HoSt) (Passing-Bablok: y = 0.97x - 0.22 ng/dL, r = 0.98, n=108) and with the Dimension Vista TTST assay (Weighted Deming: y = 1.01x - 3.32 ng/dL, r = 0.99, n=124 combined adult/pediatric; y = 1.01x - 4.74 ng/dL, r = 1.00, n=45 pediatric-only).
Interferences & pre-/post-analytical notes

Cross-reactivity

  • 5-Androstene-3β,17β-diol: -0.167% cross-reactivity (tested at 1000 ng/mL)
  • Androstenedione: 1.397% cross-reactivity (tested at 100 ng/mL)
  • Androsterone: -0.026% cross-reactivity (tested at 1000 ng/mL)
  • Canrenone: 0.066% cross-reactivity (tested at 180 ng/mL)
  • Canrenone: 0.005% cross-reactivity (tested at 500 ng/mL)
  • Canrenone: 0.008% cross-reactivity (tested at 1000 ng/mL)
  • Corticosterone: 0.014% cross-reactivity (tested at 1000 ng/mL)
  • Cortisol: 0.022% cross-reactivity (tested at 1000 ng/mL)
  • Cyproterone: 0.128% cross-reactivity (tested at 100 ng/mL)
  • Danazol: -0.121% cross-reactivity (tested at 1000 ng/mL)
  • 11-Deoxycortisol: 0.002% cross-reactivity (tested at 1000 ng/mL)
  • Dexamethasone: 0.006% cross-reactivity (tested at 1000 ng/mL)
  • DHEA: -0.042% cross-reactivity (tested at 1000 ng/mL)
  • DHEAs: 0.001% cross-reactivity (tested at 50,000 ng/mL)
  • 5α-Dihydrotestosterone: -1.410% cross-reactivity (tested at 100 ng/mL)
  • 17β-Estradiol: -0.267% cross-reactivity (tested at 1000 ng/mL)
  • Estrone: -0.009% cross-reactivity (tested at 100 ng/mL)
  • Ethisterone: 0.446% cross-reactivity (tested at 1000 ng/mL)
  • 11β-Hydroxytestosterone: 15.469% cross-reactivity (tested at 100 ng/mL) — Strong interaction; per Limitations, do not use samples from patients receiving this compound
  • 11-Keto-testosterone: 1.766% cross-reactivity (tested at 1000 ng/mL) — Strong interaction; per Limitations, do not use samples from patients receiving this compound
  • Nandrolone decanoate: N/A cross-reactivity (tested at 1000 ng/mL) — Cross-reactivity could not be calculated; compound tested above the measuring interval. Strong interaction; do not use samples from patients receiving this compound
  • Norgestrel: 0.145% cross-reactivity (tested at 1000 ng/mL)
  • Oxymetholone: -0.576% cross-reactivity (tested at 100 ng/mL)
  • Prednisolone: 0.093% cross-reactivity (tested at 1000 ng/mL)
  • Prednisone: 0.011% cross-reactivity (tested at 1000 ng/mL)
  • Progesterone: 0.014% cross-reactivity (tested at 1000 ng/mL)
  • Spironolactone: 0.105% cross-reactivity (tested at 80 ng/mL)
  • Spironolactone: 0.011% cross-reactivity (tested at 1000 ng/mL)
  • Testosterone propionate: 2.946% cross-reactivity (tested at 100 ng/mL)
  • 7α-Thiomethyl spironolactone: 0.001% cross-reactivity (tested at 400 ng/mL)
  • 7α-Thiomethyl spironolactone: -0.002% cross-reactivity (tested at 1000 ng/mL)

Interferences

Known interferences
  • Conjugated bilirubin (icterus): ≤ 10% change up to threshold; concentrations > 15 mg/dL cause erroneous results (-14.4% change demonstrated at 20 mg/dL) (15 mg/dL validated tolerance; erroneous results above this level)
  • Unconjugated bilirubin (icterus): ≤ 10% change up to threshold; concentrations > 20 mg/dL cause erroneous results (+20.1% change demonstrated at 30 mg/dL) (20 mg/dL validated tolerance; erroneous results above this level)
  • Biotin: Falsely elevated results, dose-dependent; e.g. at testosterone ~30 ng/dL, biotin at 100 ng/mL caused 4-23% bias, 250 ng/mL caused 84-107% bias, 500 ng/mL caused 370-441% bias, and ≥ 1200 ng/mL exceeded the measuring interval (≤ 10% change up to 30 ng/mL biotin; > 30 ng/mL may cause erroneous (falsely elevated) results) — Grimsey et al. 2017; Piketty et al. 2017
  • Steroid-analogue medications (e.g., Nandrolone decanoate, 11β-hydroxytestosterone, 11-keto-testosterone): Strong cross-reactive interaction causing falsely elevated results; new steroid-based medication analogues with structures similar to testosterone may also cross-react (Do not use samples from patients receiving these specific compounds)
Suspected / theoretical
  • Heterophilic antibodies (HAMA/human anti-animal antibodies): May cause falsely elevated or falsely depressed results; assay is designed to minimize (not necessarily eliminate) this interference — Kricka 1999; Vaidya & Beatty 1992
Validated -- no significant interference
  • Hemolysis (hemoglobin): ≤ 10% change in results (up to 500 mg/dL hemoglobin)
  • Lipemia (triglycerides): ≤ 10% change in results (up to 1000 mg/dL triglycerides)
  • Cholesterol: ≤ 10% change in results (up to 500 mg/dL cholesterol)
  • Rheumatoid factor: ≤ 10% change in results (up to 200 IU/mL)

Pre-analytical

  • Serum must be physically separated from cells within 2 hours of collection; do not use samples stored at room temperature > 8 hours prior to centrifugation.
  • Samples with conjugated bilirubin > 15 mg/dL or unconjugated bilirubin > 20 mg/dL will cause erroneous results.
  • Do not use samples from patients receiving Nandrolone decanoate, 11β-hydroxytestosterone, or 11-keto-testosterone due to strong cross-reactive interaction.
  • Separated specimens stable up to 48 hours at room temperature, 7 days at 2-8°C, or frozen at ≤ -20°C (up to 3 freeze/thaw cycles).

Post-analytical

  • Results > 1500.00 ng/dL (> 52.05 nmol/L) must be diluted 1:2 with Multi-Diluent 3 and retested; automatic dilution setpoint must be ≤ 1500 ng/dL (≤ 52 nmol/L).
  • Total testosterone result can be combined with an Atellica IM SHBG result to calculate the Free Androgen Index (FAI) = [Testosterone (nmol/L) / SHBG (nmol/L)] × 100, as a surrogate for physiologically active (free) testosterone.
  • Report results below the measuring interval as ≤ 7.00 ng/dL (≤ 0.24 nmol/L).
Source: package insert (PDF, Cat #10995708 (100 tests); 10995707 (500 tests), rev Rev. 07, 2023-03) · Extracted 2026-08-26 · Approved 2026-09-10
3 approved methods -- worth a cross-method comparison?

Lab comparison

No lab offerings recorded for this analyte yet.