John Kane

Macroprolactinaemia: validation and application of the polyethylene glycol precipitation test and clinical characterization of the condition

The clinical characteristics and natural history of macroprolactinaemia are yet to be fully established and there is currently no widely accepted simple in vitro test for routine identification of the condition. This study was designed to further validate the polyethylene glycol (PEG) precipitation test with assessment of percentage prolactin recovery (%R), for the detection of macroprolactinaemia and clinically characterize cases identified using the test.

A prospective study of the prevalence of clear‐cut endocrine disorders and polycystic ovaries in 350 patients presenting with hirsutism or androgenic alopecia

OBJECTIVE To determine the frequency of polycystic ovaries (PCO) on ultrasound and the incidence of clear‐cut endocrine disorders leading to virilization in patients complaining of hirsutism or androgenic alopecia. The major purpose was to determine a coherent policy for the routine biochemical assessment of such women. DESIGN A prospective study of women attending a joint skin/endocrine clinic complaining of these problems. PATIENTS Three hundred and fifty consecutive women with hirsutism and/or androgenic alopecia were assessed. MEASUREMENTS Baseline endocrine screens were conducted on two occasions and included measurement of serum testosterone, androstenedione, dehydroepiandrosterone sulphate, sex hormone binding globulin, LH, FSH, 17‐hydroxyprogesterone and PRL. The ovarles were visualized by high‐resolution pelvic ultrasound scanning. RESULTS Eight women were identified with relevant endocrine disorders; of these, one was acromegalic and one had a microprolactinoma—In both cases the association may have been fortuitous. Three had clear‐cut 21‐hydroxylase deficiency, one a rare hepatic enzyme deficiency (11‐reductase), one a virilizing adrenal carclnoma and one a Leydig cell tumour. The latter six cases all had persistently elevated levels of serum testosterone (>5nmol/l). In ail, 13 women had baseline testosterone levels in excess of 5nmol/l. Polycystic ovaries were present in 81 % of the cases who had erratic cycles and 52% of those with regular cycles; PCO were present In two of the women with 21‐hydroxylase deficiency and In the woman with 11‐oxoreductase deficiency. The Leydig cell tumour (1.2 cm diameter) was not detected on ultrasound or CT scan.

Method Comparison—A Different Approach

The commonly accepted method of analysing data from method comparison studies is regression analysis, a method which has limitations. This study illustrates the use of a graphical presentation of data, the difference plot, which can be used as an alternative to least squares regression analysis. The data from comparison studies performed on five methods were analysed both by Demings regression analysis, with calculation of the correlation coefficient, and by the difference plot. The results show that in most cases much more relevant information was obtained from the difference plot.

Analytical and clinical aspects of adrenocorticotrophin determination

Adrenocorticotrophin (ACTH) is formed from the cleavage of pro-opiomelanocortin. Measurement of plasma ACTH is a key step in the differential diagnosis of hypothalamic-pituitary-adrenal disorders. Prior to the development of radioimmunoassay, the bioassays employed for the determination of ACTH were highly complex, time-consuming and costly in terms of number of animals used. Their sensitivity was such that the normal early morning peak of ACTH could not be determined. The introduction of immunoassay methodology enabled the measurement of low normal ACTH concentrations. Immunological recognition of ACTH by the antibodies employed offered improvements with regard to specificity. The development of two-site immunometric assays further improved specificity and the ability to measure low normal ACTH concentrations without the need to extract large volumes of plasma. The quantification of ACTH is now routinely performed in clinical laboratories, with non-radioisotopic methods becoming increasingly popular. ACTH measurement is of limited value in distinguishing between the causes of Cushings syndrome, as there is considerable overlap in circulating ACTH concentrations in subjects with either a pituitary or an ectopic tumour. The role of ACTH precursor and related peptides in normal and pathological states and the clinical utility of their measurement remain to be fully elucidated. However, there is evidence that measurement of ACTH precursors can be a useful diagnostic tool in identifying the aetiology of Cushings syndrome. This review will primarily address methodological aspects of ACTH measurement in the diagnosis of clinical conditions.

Measurement of serum testosterone in women; what should we do?

All immunoassays for female serum testosterone give falsely high results in some samples. The effect is variable and cannot be predicted for any given sample. Inaccurate calibration or interference by cross-reacting substances is almost certainly the cause of the problem, but for many immunoassays, the exact nature of the interferent is not known. Some of the interference can be removed by employing an extraction step prior to immunoassay. The advent of fast simple and sensitive liquid chromatography tandem mass spectrometry methods offers an exciting alternative to immunoassay for serum testosterone measurement. It is recommended that all high serum testosterone concentrations in women are checked, before reporting, by a method which is accurate (i.e. minimal bias to isotope dilution gas chromatography mass spectrometry [ID-GCMS] method) and is not subject to interference. Action should also be taken by assay users, manufacturers, regulators and professional bodies to ensure accurate standardization and comparability of assays.

Investigation into possible causes of interference in serum testosterone measurement in women.

Background: Direct (non-extracted) testosterone immunoassays may give spuriously high results in women. The presumed interferents may be removed if testosterone is extracted into an organic phase before being measured. We aimed to clarify possible causes and effects of interference in testosterone measurement in women. Methods: Women who had a blood sample referred to Hope Hospital Clinical Biochemistry laboratory for measurement of serum testosterone concentration over a six-month period were studied. Clinical and treatment data were recorded. A difference (direct-minus-extracted testosterone) of less than 1.0 nmol/L was used to define a group with low interference. A difference of 2.5 nmol/L or more was used to define a group with significant interference. Results: The distribution of interference in 1271 serum samples from female patients was unimodal. There were no group differences in clinical presentation or treatment. The median degree of interference was 1.4 nmol/L. In 42 female patients with varying degrees of interference, identified on routine assay, regression analysis showed strong association between dehydroepiandrosterone sulphate (DHEA-S) and interference (direct-minus-extracted testosterone) (r = 0.77, P<0.001). Conclusions: Given that DHEA-S is present in very variable amounts in blood, it is possible that even with a low cross reactivity, DHEA-S or one of its metabolites significantly interferes in the testosterone assay when at higher concentrations.

Oestradiol assays: fitness for purpose?

In this review we discuss the analytical inadequacies of oestradiol assays in relation to the clinical requirements for performing them, and make recommendations for their improvement. The measurement of oestradiol can be requested in a number of clinical scenarios (precocious puberty, infertility, assisted conception, hormone replacement therapy). The very wide dynamic range of oestradiol concentrations is a huge challenge for routine assays, which they are unlikely to meet on theoretical as well as practical grounds. The EQA performance of oestradiol assays in terms of trueness, comparability, recovery and analytical sensitivity leaves much to be desired and indicates that calibration is compromised by poor analytical specificity. To make oestradiol assays fit for purpose requires concerted action by all stakeholders to define analytical quality specifications for the various clinical scenarios involved, and then to encourage concerted action by the diagnostic industry to use the steroid reference measurement system to improve specificity, trueness and traceability.

Do immunoassays differentially detect different acidity glycoforms of FSH

The possibility of the carbohydrate residues of glycoproteins affecting their recognition in immunoassays is an important and unresolved issue. This study looked for evidence of differential recognition of FSH glycoform preparations, of variable isoelectric point (pI) and known molarity, using three routine assays employing different antibody configurations.

Basal Serum Dehydroepiandrosterone Sulphate Concentration Does Not Predict the Cortisol Response to Provocative Testing

The concentration of dehydroepiandrosterone sulphate (DHEAS) was measured in the plasma of 104 patients aged 16–78 years (48 men) undergoing routine assessment of anterior pituitary reserve. A fasting plasma sample was collected at 0900 h on the same day that either an insulin tolerance test or a glucagon stimulation test was performed. Serum cortisol, DHEAS and prolactin were measured by radioimmunoassay. DHEAS levels between 0·5 and 14 nmol/L were significantly but negatively correlated with age but there was no significant correlation with basal cortisol, maximum stimulated cortisol, the increment in cortisol or basal prolactin. The correlation of DHEAS with age was also observed when results for women were analysed separately but this did not hold for men. For male and female results separately, there was again no correlation of basal DHEAS with basal cortisol, maximum stimulated cortisol, increment of cortisol or basal prolactin. The age and sex related reference range and other factors affecting basal DHEAS concentration make it a cumbersome and unreliable measure of hypothalamic-pituitary-adrenal axis reserve.

Consideration of the degree of increase in urine metadrenalines provides superior specificity in the diagnosis of phaeochromocytoma than additional urine catecholamine measurement

Background Measurement of fractionated plasma or urine metadrenalines is the recommended screening test in the diagnosis of phaeochromocytoma, with clinical cut-offs geared towards diagnostic sensitivity. Current practice at Salford Royal Hospital is to add urine catecholamines onto samples with raised urine metadrenalines, with the aim of adding specificity to a diagnosis of phaeochromocytoma. Methods This practice was reviewed by identifying a series of patients with raised urine metadrenalines who had catecholamines reflectively added. A total of 358 samples were identified from 242 patients, of which 228 had urine catecholamines measured. Results A diagnosis of ‘phaeochromocytoma‘ (n = 41) or ‘no phaeochromocytoma‘ (n = 90) was obtained in 131 of 228 patients, giving raised urine metadrenalines a positive predictive value for phaeochromocytoma of 31%. The finding of increased urine catecholamines in samples with raised urine metadrenalines increased specificity for phaeochromocytoma to 70%. However, 95% diagnostic specificity for phaeochromocytoma could be achieved by the introduction of a second cut-off for urine metadrenalines geared towards maximizing specificity. Conclusions Consideration of the degree of increase in urine metadrenalines is a superior method of determining the likelihood of phaeochromocytoma than measurement of urine catecholamines.