STEROID HORMONES

STEROID HORMONES IN SERUM LC-MS/MS KIT

Steroid hormones are defined as effective biological messengers which take part in crucial body processes even at low concentrations. Thus, reliable, and simultaneous analysis of a broad panel of steroid hormones plays vital role for the investigation of the hormone profile, which could be useful in detection endocrine disorders resulting from defects in steroid biosynthesis such as congenital adrenal hyperplasia (CAH). Due to the restrictions and shortcomings of immunoassay techniques for measurement of steroid hormones in body fluids, liquid chromatography tandem mass spectrometry (LC-MS/MS) is increasingly becoming the method for multi-class steroid hormone detection.

  • Highlights

    • Analysis of clinically relevant 16 steroid hormones in a single injection-no need for separate injections
    • Total run time is 16.0 min.
    • One sample preparation for all steroid hormones. Easy liquid-liquid extraction step following by protein precipitation for the sample treatment-no need for SPE
    • Covered corresponding isotope labeled internal standards of important steroid hormones-11 ISs
    • Long life span of HPLC column
  • Parameters

    • 17-α-Hydroxyprogesterone
    • Aldosterone
    • Testosterone
    • Estradiol
    • Progesterone
    • Cortisol
    • Dihydrotestosterone
    • Androstene-3,17-dione
    • Corticosterone
    • 11-Deoxycortisol
    • Pregnenolone
    • 11-Deoxycorticosterone
    • DHEA
    • DHEAS
    • 17-α-Hydroxypregnenolone
    • Androsterone.
  • MATRIX

    • Serum
    • Plazma

Sample Preparation for Serum


  • Step 1

    Pipette 425 µl of serum/plasma sample, calibrators or controls into a glass centrifuge tube
    (tube-1) and add 25 µl of internal standard mixture, vortex for 5 sec.

  • Step 2

    Add 450 µl of Reagent-1 into tube-1 and vortex for 15 sec. Then, centrifuge at 4000 rpm for
    5 min.

  • Step 3

    Decant the supernatant into a new glass centrifuge tube (tube-2)

  • Step 4

    Wash the precipitate in tube-1 with 1.5 mL Reagent-2 (two times!) and transfer the washing
    fluids into tube-2

  • Step 5

    Agitate tube-2 for 3 min. and centrifuge at 4000 rpm for 5 min.

  • Step 6

    Transfer 2.4 ml of the upper phase to a new glass tube (tube-3) and evaporate under
    nitrogen stream. (Upper phase can be evaporated at 7 psi during ~15 min using water bath
    at the temperature of 30 °C)

  • Step 7

    Reconstitute tube-3 with 100 µl of Reagent-3 and transfer to a HPLC vial prior to injection

  • Extracted LC-MS/MS chromatogram of Steroıd Hormones

  • Result

    Steroid Hormone

    LOQ (µg/L)

    Recoveries (%)

    Repeatability (%)

    Aldosterone

    0.035

    95.8

    6.33

    Cortisol

    0.336

    91.6

    2.48

    Corticosterone

    0.179

    102.3

    4.96

    11-Deoxycortisol

    0.048

    108.4

    3.62

    DHEAS

    16.32

    105.9

    2.97

    Estradiol

    0.051

    108.7

    5.99

    Testosterone

    0.026

    93.4

    1.88

    DHEA

    0.336

    109.2

    8.57

    11-Deoxycorticosterone

    0.052

    95.8

    7.73

    Androstene-3,17-dione

    0.076

    97.4

    2.47

    17-α-Hydroxyprogesterone

    0.091

    97.2

    4.31

    Dihydrotestosterone

    0.041

    104.0

    9.92

    Androsterone

    0.115

    87.8

    9.39

    Pregnenolone

    0.108

    92.4

    7.53

    17-α-Hydroxypregnenolone

    0.480

    99.3

    6.55

    Progesterone

    0.108

    113.9

    7.11

ADAPTING DIAGNOSTICS IN CHROMATOGRAPHY COUPLED MASS SPECTROMETRY..

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