A history of clinical biochemistry through the eye of an enzyme

The Royal College of Pathologists recently published a book entitled History of Pathology in 50 Objects. ¹ This approach reflects a vogue for using often simple objects as a lens through which to study social history. I have attempted a similar process using the now very workaday analyte alkaline phosphatase (ALP) to provide a perspective on the history of clinical biochemistry. A review of its appearances in the Annals, sometimes as the star player in a paper and sometimes just as a bit part in a description of an altogether different topic has proved fascinating.

The presence of phosphatase activity in bone was first described by Robert Robison in 1923 ^:2 by 1930, it had been measured in blood and found to be elevated in bone disease and obstructive liver disease. The first analytical method for clinical use was developed by Earl King and Riley Armstrong in 1934. ³ During this period, ALP was used primarily as a marker of bone disease although it was recognized that its activity was also increased in liver disease.

ALP made its first appearance in the very first volume of the Annals, in a report of a lecture given in May 1961 at a meeting of the ACB Midland Region, by Noel Maclagan, charmingly entitled ‘A comparison of the efficacy of some popular tests in diagnosis’. ⁴ Maclagan quoted data from The Westminster Hospital demonstrating that ALP performed as efficiently as the thymol turbidity test in the diagnosis of obstructive liver disease. In the same volume, there was a proposal for standardization of enzyme units based on µmol of substrate consumed, ⁵ yet 25 years later, the Annals was still publishing papers quoting ALP in King-Armstrong units.

The early 1960s also saw the development of electrophoretic methods for the separation of ALP isoenzymes (described by Albert Latner in 1961), ⁶ and further characterization of its enzymatic properties by Donald Moss. Moss published a detailed account of urinary ALP in 1964, ⁷ only to conclude that it ‘does not appear to offer much diagnostic value’ – how many researchers would be this candid today?

By this time, clinical enzymology was all the rage. Denis Barron described it as ‘the new magic’ but bemoaned that fact that his laboratory at The Royal Free Hospital had measured ALP in >4000 samples in 1964. ⁸ Help was at hand, however, with the development of the automated analyser; Anthony Pollard outlined the principals of the early techniques (largely continuous flow) ⁹ and a 1967 paper ¹⁰ described a methodology that could process as many as 50 samples per hour. As an aside, the advent of automation led to reorganization of services into ‘cold’ and ‘hot’ labs to take advantage of economies of scale (ALP was a ‘cold’ laboratory assay). ¹¹ Even using automated analysers, many scientists were still making up their own reagents, as described in a technical bulletin on kit methods co-authored by another of the great clinical enzymologists, Sidney Rosalki, in 1972. ¹²

The development of external quality assessment schemes was another feature of the 1970s. Reports of these^13,14 make interesting reading not just because of significant between-laboratory variation but also because they list the wide range of manual and autoanalyser methods still in use. Publication of the International Federation of Clinical Chemistry recommended method in 1983 ^{1 5} consolidated a shift towards the use of 4-nitrophenylphosphate as substrate, with a phosphate-accepting buffer (although 15 years later, some 30% of UK laboratories continued to use the alternative Scandinavian/German method) ¹⁶ and Moss, in one of his last publications in 1997, ¹⁷ proposed that the use of enzyme calibrators could enable several well-proven recommended methods to remain in use.

Alkaline phosphatase first appeared in the immunoassay arena as one of the growing number of non-isotopic labels, as described in 1979. ¹⁸ Shortly thereafter, ALP measurement by immunoassay became possible. Chris Price, in a 1993 review of isoenzyme methods (including lectin precipitation and chemical and heat inactivation), ¹⁹ described the challenge of producing a monoclonal antibody specific to bone ALP, and from 1994 onwards, the Journal contains a significant number of papers describing the clinical applications of bone ALP immunoassays.

The 21st century could be called ‘ALP, the mature years’, a period marked by a reduction in the number of Annals papers focusing on new developments. There is however still a smattering of interesting papers describing novel isoforms, the most recent appearing in 2012. ²⁰ It is good to see that this enzyme still interests clinical biochemists today, but the heady days of Barron’s ‘new magic’, when many of the old academic departments were led by enzymologists who could claim an almost apostolic line of descent from Earl King, are well and truly gone.