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Salivary Testosterone ELISA Kit


Technical Summary

Assay Protocol
Rev. 09.09.22
Catalog#: 1-2402
Regulatory Status: RUO
Format: 96-well plate
Assay Time: ~ 2 hrs
Sample Volume/Test: 25 µL
Sensitivity: 1 pg/mL
Assay Range: 6.1 pg/mL - 600 pg/mL
Storage Requirements: 2-8°C
Tests Per Kit
Singlet: 76
Duplicate: 38
Target Analyte
Technical Documentation

Salivary Testosterone Assay Kit Overview

Intended Use

The Salimetrics Testosterone Enzyme Immunoassay Kit is a competitive immunoassay specifically designed and validated for the quantitative measurement of salivary Testosterone. It is not intended for diagnostic use. It is intended only for research use in humans and some animals. Salimetrics has not validated this kit for serum or plasma samples.


Testosterone is an anabolic steroid hormone synthesized from androstenedione in the Leydig cells of the testes of males and, in smaller quantities, in the ovaries of females. Small amounts are also secreted by the adrenal glands in both sexes. Testosterone production also occurs in peripheral tissues by conversion of circulating DHEA-S, DHEA, and androstenedione. Testosterone exhibits a diurnal rhythm, with highest levels in the morning and a nadir around midnight. In men, Testosterone plays an important role in the development of male reproductive tissues including the testes and prostate, as well as promoting secondary sexual characteristics such as increased muscle, bone mass, and hair growth. In blood, only 1-10% of Testosterone is in its unbound or biologically active form. The remaining Testosterone is bound to serum proteins. Unbound Testosterone enters saliva via intracellular mechanisms, and in saliva the majority of Testosterone is not protein-bound. Salivary Testosterone levels are unaffected by salivary flow rate. The serum-saliva correlation for Testosterone is very high for males, but only modest for females, possibly because women’s values often fall near the bottom of the measurable range for both serum and saliva immunoassay kits.

Salivary Testosterone Assay Principle

This is a competitive immunoassay kit. Testosterone in standards and samples compete with Testosterone conjugated to horseradish peroxidase for the antibody binding sites on a microtitre plate. After incubation, unbound components are washed away. Bound Testosterone Enzyme Conjugate is measured by the reaction of the horseradish peroxidase enzyme to the substrate tetramethylbenzidine (TMB). This reaction produces a blue color. A yellow color is formed after stopping the reaction with an acidic solution. The optical density is read on a standard plate reader at 450 nm. The amount of Testosterone Enzyme Conjugate detected is inversely proportional to the amount of Testosterone present in the sample.

References & Salivary Testosterone Research

    1. Labrie, F., Luu-The, V., Bélanger, A., et al. (2005).  Is dehydroepiandrosterone a hormone? J Endocrinol, 187(2), 169-96.
    2. Nakamura, Y., Hornsby, P.J., Casson, P., et al. (2008).  Type 5 17β-hydroxysteroid dehydrogenase (AKR1C3) contributes to testosterone production in the adrenal reticularis. J Clin Endocrinol Metab, 94(6), 2192-98.
    3. Burger, H.G. (2002). Androgen production in women. Fertil Steril, 77(Suppl 4), S3-5.
    4. Labrie, F., Bélanger, A., Cusan, L., Candas, B. (1997).  Physiological changes in dehydroepiandrosterone are not reflected by serum levels of active androgens and estrogens but of their metabolites: Intracrinology.  J Clin Endocrinol Metab, 82(8), 2403-9.
    5. Ankarberg, C., Norjavaara, E. (1999).  Diurnal rhythm of testosterone secretion before and throughout puberty in healthy girls: Correlation with 17β-estradiol and dehydroepiandrosterone sulfate.  J Clin Endocrinol Metab, 84(3), 975-84.
    6. Diver, M.J., Imtiaz, K.E., Ahmad, A.M., et al. (2003).  Diurnal rhythms of serum total, free and bioavailable testosterone and of SHBG in middle-aged men compared with those in young men. Clin Endocrinol (Oxf), 58(6), 710-17.
    7. Rogol, A.D., Clark, P.A., Roemmich, J.N. (2000).  Growth and pubertal development in children and adolescents: Effects of diet and physical activity.  Am J Clin Nutr, 72(2 Suppl.), 521S-28.
    8. Snyder, P.J., Peachey, H., Berlin, J.A., et al. (2000). Effects of testosterone replacement in hypogonadal men.  J Clin Endocinol Metab, 85(8), 2670-77.
    9. Tibblin, G., Adlerberth, A., Lindstedt, G., Björntorp, P. (1996).  The pituitary-gonadal axis and health in elderly men: A study of men born in 1913.  Diabetes, 45(11), 1605-9.
    10. Davis, S.R., Tran, J. (2001). Testosterone influences libido and well being in women. Trends Endocrinol Metab, 12(1), 33-7.

    1. Wang, C., Alexander, G., Berman, N., et al. (1996). Testosterone replacement therapy improves mood in hypogonadal men: A clinical research center study. J Clin Endocrinol Metab, 81(10), 3578-83.
    2. Malkin, C.J., Pugh, P.J., West, J.N., et al. (2006). Testosterone therapy in men with moderate severity heart failure: A double-blind randomized placebo controlled trial.  Eur Heart J, 27(1), 57-64.
    3. Bhasin, S., Bremner, W.J. (1997). Clinical review 85: Emerging issues in androgen replacement therapy.  J Clin Endocrinol Metab, 82(1), 3-8.
    4. Gibson, M., Lackritz, R., Schiff, I., Tulchinsky, D. (1980). Abnormal adrenal responses to adrenocorticotropic hormone in hyperandrogenic women.  Fertil Steril, 33(1), 43-8.
    5. Rodin, A., Thakkar, H., Taylor, N., Clayton, R. (1994). Hyperandrogenism in polycystic ovary syndrome: Evidence of dysregulation of 11β-hydroxysteroid dehydrogenase.  N Eng J Med, 330(7), 460-65.
    6. Vining, R.F., MicGinley, R.A. (1987). The measurement of hormones in saliva: Possibilities and pitfalls.  J Steroid Biochem, 27(1-3), 81-94.
    7. Wang, C., Plymate, S., Nieschlag, E., Paulsen, C.A. (1981). Salivary testosterone in men: Further evidence of a direct correlation with free serum testosterone.  J Clin Endocrinol Metab, 53(5), 1021-24.
    8. Rollin, G. (2010). The trials of testosterone testing.  Clin Lab News, 36(8), 1-5.

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