Immunoradiometric method and electrophoretic system compared for quantifying bone alkaline phosphatase in serum.

Agarose electrophoresis (Isopal, Beckman) and an immunoradiometric assay (IRMA) involving specific monoclonal antibodies (Ostase, Hybritech), two methods for the quantification of serum bone alkaline phosphatase (ALP, EC 3.1.3.1), a marker of osteoblastic activity, were compared in 293 patients: 79 with end-stage renal failure treated with hemodialysis and 214 with malignant disease. Overall correlation between the two methods was good (r = 0.92), except (a) for low values of bone ALP and (b) in some samples with high total liver ALP activity--both due to considerable cross-reactivity of the anti-bone ALP antibodies of the Ostase kit with liver ALP. This interference was not constant and was not evenly distributed across all concentrations of bone ALP. Low bone ALP determined with the IRMA (< or = 5 micrograms/L) was confirmed by electrophoresis (< or = 21 U/L), but bone ALP activity determined by electrophoresis to be low (< or = 21 U/L) was not correlated with the IRMA results. After standardizing our results by computing z-values for bone ALP, delta z (= zOstase - zIsopal) was significantly correlated with liver ALP activity (r = 0.73, P < 0.0001). We conclude that the IRMA for quantifying bone ALP is acceptable as a screening method. However, when high values for bone ALP are found with the Ostase method, confirmation by electrophoresis remains mandatory to rule out cross-reactivity with high amounts of liver ALP. For detecting low bone ALP activities, electrophoresis remains the method of choice.

Alkaline phosphatase isoenzyme patterns in malignant disease.

Early treatment of patients with malignant disease and liver or bone metastasis may increase their survival time. We have used the activity patterns of liver and bone isoenzymes of alkaline phosphatase (ALP), separated by agarose gel electrophoresis, to detect early metastasis. We studied ALP isoenzyme patterns in a background population of 101 patients with no evidence of any disease that might influence this pattern; a healthy reference population (n = 330); and the following three groups of patients: 143 with malignant disease, 47 with nonmalignant liver disease, and 22 with nonmalignant bone disease. Cutoff and predictive values of liver ALP, high-molecular-mass (high-M(r)) ALP, and bone ALP were established for detecting liver and bone metastasis. The positive predictive value of liver and high-M(r) ALP was higher than that of total ALP in detecting liver metastasis, but liver and high-M(r) ALP did not enable us to differentiate between malignant and nonmalignant liver disease. Total ALP activity was of slightly more value than liver and high-M(r) ALP in enabling us to rule out liver metastasis. From bone ALP activity we could not distinguish between nonmalignant bone disease and bone metastasis. The negative predictive value of bone ALP in the diagnosis of bone metastasis was low, but its positive predictive value was high and superior to that of total ALP.

Age and sex distribution of alkaline phosphatase isoenzymes by agarose electrophoresis.

We separated isoenzymes of alkaline phosphatase (ALP; EC 3.1.3.1) in 1383 sera of normal individuals (ages 4-65 years) by agarose electrophoresis with the Isopal system (Analis). As expected, the predominant isoenzyme in children was of bone origin, and almost all (99%) of the children had low activities of a second bone fraction, "bone variant" ALP. The "bone variant" disappeared after age 17 in girls and after age 20 in boys. The highest (median) bone ALP activity was reached at age 9 to 10 in girls and at age 13 to 14 in boys, followed by a gradual decline in girls and a steep decline in boys. During adulthood, activity of the bone fraction was constant and no significant differences were observed between sexes, neither for bone nor for liver ALP activity. The latter remained almost unchanged throughout life. We observed no high-Mr ALP activity in children, whereas sera from 60% of the adults contained low activities of high-Mr ALP. Intestinal ALP (soluble form) and "intestinal variant" ALP (hydrophobic form) were frequently present, in 21% and 37% of all samples, respectively. No significant differences were observed between age groups and sexes for the intestinal isoenzymes.

Improved agarose electrophoretic method for separating alkaline phosphatase isoenzymes in serum.

A modified agarose electrophoretic system for the separation of alkaline phosphatase (ALP, EC 3.1.3.1) isoenzymes is described. Bone, liver, high-molecular-mass, and intestinal ALP are separated with high reproducibility. The sensitivity of the agarose system is superior to cellulose acetate in detecting high-Mr ALP. Correlation is good between bone ALP fractions scanned before and after treatment with neuraminidase. Immunoglobulin-bound ALPs, the ALP-lipoprotein-X complex, and the additional ALP fraction observed in transient hyperphosphatasemia in children are detected by their peculiar electrophoretic mobility in the proposed system. Approximately 25% of the samples contained an additional fraction of intestinal-type ALP, as evidenced by neuraminidase treatment and use of polyclonal and monoclonal antibodies. Because the electrophoretic mobilities of this "intestinal variant" and of some immunoglobulin-bound ALP fractions are identical to those of bone and intestinal ALP, respectively, treatment of the samples with a polyclonal antibody that reacts with intestinal ALP is advised.