One Small Draw, the Whole Endocrine Map

1. Background

Hormones do not act in isolation. Adrenal and gonadal steroids share a common biosynthetic tree; binding proteins set what fraction is bioavailable; pituitary signals drive the peripheral glands; and metabolic and vitamin-D status modulate the whole.

Testing that samples one hormone at a time, on separate venous draws, sees the network only in fragments.

This review describes an alternative made practical by two technologies — multiplexed LC-MS/MS measurement and small-volume microsampling — in which a single capillary collection maps the endocrine network broadly enough to read not just individual hormones but the relationships among them.

2. One collection, many analytes

Multiplexed LC-MS/MS assays routinely quantify large steroid panels from a single specimen: published methods measure fifteen gluco- and mineralocorticoid-pathway steroids in one run, and thirteen steroids simultaneously in clinical cohorts.^2,3

The same chemistry works from very small volumes. A validated method quantifies nine steroids — cortisol, 17-hydroxyprogesterone, 11-deoxycortisol, 21-deoxycortisol, androstenedione, corticosterone, 11-deoxycorticosterone, testosterone and progesterone — from a microsample.¹

Clinicore’s capillary panel runs 29 analytes from a single ~600 µL collection: the 20 LC-MS/MS endocrine targets, plus nine protein hormones and markers by immunoassay. Why those two platforms sit on one collection is set out in §4.3.

Collection routes, volumes and stability are on the microsampling page.

3. The steroidogenic tree

Most of the panel’s steroids are nodes on a single biosynthetic tree running from cholesterol through pregnenolone and progesterone into four branches:

• Mineralocorticoids — deoxycorticosterone, corticosterone, aldosterone
• Glucocorticoids — 11-deoxycortisol, cortisol, and its inactive partner cortisone
• Androgens — DHEA, androstenedione, testosterone, DHT
• Estrogens — estrone, estradiol, estriol

Because the panel measures precursors and products together, the ratio of a product to its precursor reports the activity of the enzyme between them — the principle behind reading 21-hydroxylase from 17-hydroxyprogesterone in congenital adrenal hyperplasia.

A single multiplexed panel therefore does more than list hormones; it exposes the enzymatic structure of the pathway.

4. Reading the panel, axis by axis

4.1 Adrenal and corticosteroid pathway

Cortisol, cortisone, corticosterone, aldosterone, 11-deoxycorticosterone, 11-deoxycortisol, 21-deoxycortisol, 17-hydroxyprogesterone, pregnenolone and progesterone together cover the adrenal tree.

The cortisol-to-cortisone ratio indexes 11β-HSD2 activity; 17-hydroxyprogesterone with 21-deoxycortisol flags 21-hydroxylase deficiency; and multi-steroid profiling subtypes adrenal disorders by pattern rather than by any single value.

4.2 Sex steroids

DHEA, DHEA-S, androstenedione, testosterone, DHT, estrone, estradiol and estriol map the sex-steroid pathway. The panel supports the diagnostic ratios developed across this series — testosterone:androstenedione for 17β-HSD, DHT:testosterone for 5α-reductase, and estradiol:estrone for aromatisation and menopausal status.

Each of those ratios has a measuring-interval constraint stated in its own review: see the androgen and estrogen reviews.

4.3 Binding protein, gonadotropins and metabolic context

Nine further analytes are validated on the same collection: SHBG, the gonadotropins FSH and LH, prolactin, TSH, growth hormone, insulin, and PSA total and free.

SHBG matters because it sets bioavailability — with total testosterone it gives calculated free testosterone and the free androgen index, which reflect the bioavailable fraction more accurately than a total alone. The free testosterone calculator computes it. The gonadotropins place the peripheral steroids in pituitary context, and insulin ties the panel to the metabolic dimension.

Two platforms from one collection, by design

These nine run on immunoassay, not LC-MS/MS. That is not a compromise in the panel, and it is worth saying why rather than letting the reader assume one method throughout.

Steroids are small molecules of near-identical structure. Cortisol and cortisone differ by a single oxidation state; 21-deoxycortisol, 11-deoxycortisol and corticosterone are isobaric. An antibody raised against one binds the others, and the reported concentration becomes the sum of the target and its look-alikes. That is the problem LC-MS/MS exists to solve, and Methods M1 sets out what it costs when it is not solved.

Protein hormones are not like that. SHBG, the gonadotropins, prolactin, TSH, growth hormone and insulin are large molecules with distinct surfaces. An antibody raised against one binds that one. The cross-reactivity that makes immunoassay unreliable for steroids does not apply to them, and no structural method is needed to get an accurate answer.

So the panel is two-tier because the analytes are two kinds of molecule. Mass spectrometry where the question demands it; a cleared immunoassay where a good one exists and the chemistry does not require more. A laboratory claiming every assay on its menu is novel would not be describing engineering judgment.

Every analyte page on this site states which platform measures it.

What the capillary collection does not carry

The thyroid hormones are venous only. Free and total T3 and T4 are not on the capillary panel and require a venous draw. They are iodothyronines — small molecules rather than protein hormones — and the reasoning above does not extend to them.

TSH is on the capillary panel; the thyroid hormones it regulates are not. Where a thyroid workup is the clinical question, plan a venous collection.

4.4 Vitamin D status

25-hydroxyvitamin D is the accepted marker of vitamin-D status. Note the scope on the 25-OH vitamin D3 page: this method measures D3, so a patient supplementing with ergocalciferol carries D2 the method does not capture.

5. The ratios that matter

Because the panel measures the pathway rather than isolated points, a set of clinically meaningful ratios can be computed from a single collection. Each reads either an enzyme step or a physiologic balance.

Ratio	Reads	Typical context
Cortisol : cortisone	11β-HSD2 activity	Mineralocorticoid hypertension; free-cortisol status
17-OHP (± 21-deoxycortisol)	21-hydroxylase	Congenital adrenal hyperplasia; androgen excess
11-deoxycortisol : cortisol	11β-hydroxylase	CAH; mineralocorticoid hypertension
Corticosterone → aldosterone	Aldosterone synthase	Mineralocorticoid pathway
DHEA : DHEA-S	Active : reservoir	Adrenal androgen dynamics
Testosterone : androstenedione	17β-HSD activity	Androgen production
DHT : testosterone	5α-reductase activity	Peripheral androgen activation
Estradiol : estrone	Aromatisation, menopause, route	Estrogen status
Cortisol : DHEA	HPA balance	Stress; adrenal reserve

Every ratio here depends on each analyte being measured accurately. That is the argument of Methods M1, and it is the precondition for this entire panel: a cortisol-to-cortisone ratio is meaningless if the cortisol antibody is also counting cortisone.

6. Why one small-volume collection changes the picture

Three properties compound.

Breadth. Measuring the network together makes the ratios above available from one specimen, rather than reconstructing them across separate orders drawn on different days.

Specificity and sensitivity. LC-MS/MS resolves structurally similar steroids that cross-react on immunoassay, and remains accurate at low concentrations.

Access. A collection that does not require a phlebotomist makes timed and serial sampling practical — which matters most for the analytes whose value depends on when they were drawn.