The Leeds Teaching Hospitals NHS Trust


Useful Information

Hormone Assay Interference

Interference in immunoassay is a well recognised problem. This may be due to a number of factors such as non-specificity of antibodies used in the assay method or Interference from a component in the patient sample such as circulating antibodies. These interfering antibodies are specific to an individual patient and have the potential to interfere in an unpredictable way. The reported prevalence of such interfering antibodies varies from 0.05 to more than 2%.  Our experience with thyroid hormones and gonadotrophins is that interference occurs in approximately 0.5% clinical samples.

The approach to detection of interference that we have taken is:

1: alertness to the problem

2: to use an alternative analytical method either a different immunoassay method, or by organic extraction prior to analysis or by mass spectrometry.

3: to use PEG precipitation to remove interfering proteins

4: to use commercially available anti-animal antibody coated tubes to remove any endogenous antibody


Ismail AAA, J.H.Barth JH, Walker PL, Cawood M. Interference in immunoassay is an underestimated problem. Ann Clin Biochem 2002;39:366-373

Ismail AAA, Walker PL, Barth JH, Lewandowski KC, Jones RG, Burr WA. Wrong biochemistry results: two case reports and observational study in 5310 patients on potentially misleading thyroid-stimulating hormone and gonadotropin immunoassay results. Clin Chem 2002;48:2023-2029

Ismail AAA, Walker PL, Fahie-Wilson MN, Jassam N, Barth JH. Prolactin and macroprolactin: a case report of hyperprolactinaemia highlighting the interpretation of discrepant results. Ann Clin Biochem 2003;40:298-300

Gynaecological Endocrinology

Day 21 progesterone (nmol/L) for evaluation of the function of the corpus luteum:

Day 21 progesterone is a misnomer as it is only correct for women with 28 day cycles. In order to assess optimal luteal function, progesterone measurements should ideally be made 7 days prior to the next menstrual bleed.


poor luteal function: ovulation unlikely


optimal luteal function indicating ovulation likely


may indicate suboptimal luteal function unless there is multiple ovulation due to either spontaneous occurrence or due to induction by clomiphene





Premature ovarian insufficiency

both high but usually FSH > LH


Weight loss associated amenorrhoea



Polycystic ovary syndrome

LH: FSH ratio is > 2.5:1 in many cases but is an unreliable test for the diagnosis of PCO



low LH and FSH


Oestrogen-secreting tumour



Prolactinoma (see below)




Mildly elevated values ie 600-900 U/L may be due to the stress of venepuncture; samples with values in this range should be checked on a repeat sample.

Elevated levels of prolactin may be due to pregnancy, hypothyroidism and drugs eg phenothiazines, haloperidol, tricyclic anti-depressants, metoclopramide, methyl DOPA & high dose oral contraceptives.

Elevated prolactin levels in the absence of the above conditions requires further investigation for prolactinoma eg full pituitary function tests and imaging.

Biochemical changes during pregnancy

Real changes

§  elevation in hCG

§  increase in alkaline phosphatase due to placental production of the isoenzyme

§  decrease in albumin, creatinine and urea

Apparent changes

§  There is an oestrogen-induced increase in several plasma binding proteins which apparently increases the levels of protein bound substances although their activity is unchanged as the free levels are unchanged. This may result in an elevation in plasma thyroxine, T4 > 200 nmol/L.

Investigation of the hirsute woman

Clinical evaluation

First line investigations

Second line investigations

long-standing hirsuties, regular menstrual cycles, no virilism

none necessary


long-standing hirsuties, irregular menstrual cycles, no virilism

investigate for PCO (ovarian ultra-sound or LH: FSH ratio). Testosterone >5.5 nmol/L

if Testosterone > 5.5 nmol/L proceed as below for CAH or tumour

severe hirsuties, irregular menstrual cycles and virilism

investigate for CAH if long history and for adrenal or ovarian androgen-secreting tumour if history is short