Intestinal Alkaline Phosphatase Isoenzymes Research Articles

The role of intestinal alkaline phosphatase in the development of obesity. Modulation of enzyme activity by high fat diet and dietary fiber

Interest to the tissue-specific intestinal isoenzyme of alkaline phosphatase (IAP) has increased in recent years due to eating disorders that have led to widespread obesity and diet-related diseases. Obesity is considered as an inflammation of low intensity, which is accompanied by the manifestation of various metabolic complications and a disturbance of intestinal homeostasis. IAP is one of the participants in the mechanism of the macroorganism protection against inflammatory and infectious processes, carrying out enzymatic detoxification of bacterial lipopolysaccharide (the trigger of the inflammatory process). Deficiency of IAP activity contributes to the risk of obesity, inflammatory diseases. The objective of the research was to summarize the current understanding of the role of IAP involved in the molecular mechanism of diet-induced obesity and to evaluate the impact of dietary components - fats and dietary fiber on IAP activity. Material and methods. A literature search on the role of IAP in the development of obesity was carried out using PubMed, Scopus, Web of Science, Google Scholar, ResearchGate, RSCI databases. Results. IAP prevents the development of the inflammatory process by participating in the detoxification of toxic bacterial products, limiting the translocation of pathogenic bacteria from the intestine to various tissues and organs of the macroorganism. The enzyme maintains the integrity of the intestinal barrier, influencing the synthesis and proper localization of tight junction's proteins between intestinal epithelial cells, promotes changes in the composition of the microbiota, decreasing pathogenic bacteria and increasing the population of the community of beneficial microorganisms. IAP is involved in the regulation of fatty acid absorption and influences on the adipogenesis. Monitoring the activity of IAP present in human stool can predict the early development of such complications associated with obesity as metabolic syndrome and diabetes mellitus, Some nutrients modulate IAP activity. Depending on the amount, type, composition of fats and the duration of their consumption, either an increase or decrease in the IAP activity are observed, while dietary fibers stimulate the activity of the enzyme. Conclusion. IAP activity can be considered as an early predictor of the risk of obesity. Deficiency of IAP activity contributes to the development of obesity caused by high-fat diet. The high activity of the enzyme contributes to the support of intestinal homeostasis and limits transepithelial movement of bacteria, weakening the inflammatory process induced by lipopolysaccharides, the excess concentration of which is detected in obesity. Stimulating enzyme activity through dietary intervention reduces the risk of obesity and metabolic complications.

Comment on “Relationship of nonvisualization of the fetal gallbladder and amniotic fluid digestive enzymes analysis to outcome”

Objective The aims of this study were evaluate the significance of non-visualization of fetal gallbladder at routine ultrasound scan in a series of 102 cases and to determine the contribution of amniotic fluid digestive enzyme (AF-DE) analysis towards the outcome. Method This is a multicenter retrospective study. Outcome of pregnancies, karyotype, and result of screening for CFTR gene mutations were known in all cases. Amniotic fluid gamma-glutamyl-transpeptidase and intestinal alkaline phosphatase isoenzyme were assayed. Results Non-visualization of the fetal gallbladder was associated with a severe disease in 25 cases (cystic fibrosis in ten, biliary duct atresia in eight, digestive tract anomalies in six, and chromosomal anomaly in one). In the remaining 77 cases, gallbladder agenesis was diagnosed in 22, and in 55, the gallbladder was subsequently demonstrated. Before 22 weeks of gestation (n = 30), an abnormal AF-DE pattern had a 90% sensitivity and 80% specificity in detecting cystic fibrosis or biliary duct atresia. After 22 weeks, sensitivity fell to 53%. The AF-DE pattern was normal in 82% of gallbladder agenesis cases (benign) and in 91% of the cases where the gallbladder was subsequently detected. Conclusion Non-visualization of the fetal gallbladder was associated with severe anomalies in 24% of cases. Prior to 22 weeks, determination of AF-DE contributes to the prediction of biliary atresia or the presence of cystic fibrosis. © 2012 John Wiley & Sons, Ltd.

Biochemical analysis of cystic fluid in the diagnosis of fetal intra‐abdominal masses

The aim of the study was to evaluate a biochemical analysis of fluid sampled in utero from fetal abdominal cystic masses. A retrospective study of 42 intra-abdominal cystic masses [ovarian cysts (n = 22), cloaca (n = 8), urodigestive fistulae (n = 4), and urogenital sinuses (n = 8)] was carried out. The control group (n = 30) consisted of fetal urine. Seven biochemical markers were assayed: sodium, estradiol, β(2) -microglobulin, total proteins and digestive enzymes, gamma-glutamyl transpeptidase, aminopeptidase M, and intestinal alkaline phosphatase isoenzyme. Outcome of pregnancies and final diagnosis of intra-abdominal mass were known in all cases. Biochemical patterns allowed to demonstrate (1) an ovarian origin based on elevated level of estradiol and of total proteins (100% specificity, 100% sensitivity); (2) urodigestive fistula based on the presence of high levels of digestive enzymes (cloaca in female fetuses or recto-urethral fistula in male fetuses); and (3) a renal origin (urinary pattern with low total proteins and absence of digestive enzymes); however, a biochemical pattern could not differentiate between fetal urogenital sinus and megacystis. Etiological diagnosis of a fetal cystic abdominal mass could impact both prenatal and postnatal management providing adapted prenatal counseling by a pediatric surgeon in surgically correctable congenital anomalies.

Serum alkaline phosphatase isoenzymes in SD rats detected by polyacrylamide-gel disk electrophoresis

Serum alkaline phosphatase (ALP) activity is frequently measured in toxicity studies. In the present study, we assessed the usefulness of a commercially available polyacrylamide-gel disk electrophoresis kit used in humans (AlkPhor System, Jokoh Co. Ltd., Tokyo, Japan) for identifying serum ALP isoenzymes in rats of the Sprague-Dawley strain (SD rats), which are commonly used in toxicity studies. We also examined age-related changes in serum ALP isoenzymes in SD rats. In order to identify the origin of each ALP isoenzyme, tissue ALP extracts from the liver, bone and small intestine (SI) and serum samples were treated with neuraminidase, antiintestinal ALP antibody, ALP inhibitor levamisole, and/or wheat germ agglutinin. It became clear that pretreatment of serum with neuraminidase is necessary for rat serum ALP isoenzyme analysis. The kit revealed that the main serum ALP isoenzymes in fasted 8-week-old intact rats were bone- and SI-derived and they tended to decrease with age. Serum liver-derived isoenzyme was slightly detected in both sexes of all ages examined, but it greatly increased in cholestasis model rats with bile-duct ligation, and rats of this model also had large molecular ALP detected in the stacking gel, suggesting hepatic damage. High-molecular intestinal ALP isoenzyme was slightly observed at the most cathodal side of the resolving gel. These results suggest that the present method is a useful tool for detecting serum ALP isoenzymes in SD rats and that concomitant levamisole inhibition with another gel is applicable for the evaluation of organ toxicity.

Relationship of non‐visualization of the fetal gallbladder and amniotic fluid digestive enzymes analysis to outcome

The aims of this study were evaluate the significance of non-visualization of fetal gallbladder at routine ultrasound scan in a series of 102 cases and to determine the contribution of amniotic fluid digestive enzyme (AF-DE) analysis towards the outcome. This is a multicenter retrospective study. Outcome of pregnancies, karyotype, and result of screening for CFTR gene mutations were known in all cases. Amniotic fluid gamma-glutamyl-transpeptidase and intestinal alkaline phosphatase isoenzyme were assayed. Non-visualization of the fetal gallbladder was associated with a severe disease in 25 cases (cystic fibrosis in ten, biliary duct atresia in eight, digestive tract anomalies in six, and chromosomal anomaly in one). In the remaining 77 cases, gallbladder agenesis was diagnosed in 22, and in 55, the gallbladder was subsequently demonstrated. Before 22 weeks of gestation (n=30), an abnormal AF-DE pattern had a 90% sensitivity and 80% specificity in detecting cystic fibrosis or biliary duct atresia. After 22 weeks, sensitivity fell to 53%. The AF-DE pattern was normal in 82% of gallbladder agenesis cases (benign) and in 91% of the cases where the gallbladder was subsequently detected. Non-visualization of the fetal gallbladder was associated with severe anomalies in 24% of cases. Prior to 22 weeks, determination of AF-DE contributes to the prediction of biliary atresia or the presence of cystic fibrosis.

Alkaline Phosphatases From Camel Small Intestine

Camel intestinal alkaline phosphatase have been purified and characterized. The purification was carried out by chromatography on DEAE-cellulose. Five intestinal alkaline phosphatase isoenzymes (IAP1 to IAP5) were obtained. IAP2 and IAP5 with the highest activity levels were purified to homogeneity by Sephacryl S-200 column. The molecular weights of camel small intestinal IAP2 and IAP5 were estimated to be 36,000 Da and 49,800 Da, respectively using Sephacryl S-200. These values were confirmed by SDS-PAGE, where the two enzymes migrated on the gel as single subunits and the molecular weights were 35,000 Da and 49,000 Da, respectively. IAP2 and IAP5 had isoelectric points at 5.4 and 5.7 and Km values of IAP2 and IAP5 are 0.26 and 0.34 mol p-nitrophenyl phosphate/ml, respectively. The two alkaline phosphatases IAP2 and IAP5 were found to have common character that they had higher affinity toward p-nitrophenyl phosphatae and α-naphthyl phosphate compared to the other studied substrates. IAP2 and IAP5 had pH optima at 10 and 10.5 and temperature optima at 50ºC and 40ºC with activation energy 9.3 and 9.6 Kcal mol-1, respectively. The two isoenzymes were stable up to 40ºC and 50ºC, respectively. The effect of different metal cations on the activity of camel intestinal IAP2 and IAP5 was studied. The camel intestinal alkaline phosphatase isoenzymes IAP2 and IAP5 were inhibited by EDTA and phenylalanine. Keywords: Camel; Small intestine; Alkaline phosphatase ; Purification; Characterization Egyptian Journal of Biochemistry and Molecular Biology Vol. 26 (1) 2008 pp. 31-54

Elevated serum alkaline phosphatase levels in a renal transplant patient precede colitis

An asymptomatic, but highly significant, rise in serum alkaline phosphatase (AP) levels developed in a renal transplant recipient. Investigations ruled out bony or hepatobiliary disease. Subsequent diarrhea and weight loss led to a diagnosis of cytomegalovirus (CMV) colitis, which was confirmed with a positive CMV pp65 antigenemia test and an endoscopic finding of multiple colonic erosions. Intravenous ganciclovir led to complete patient recovery and a swift reduction of serum AP levels to normal. Normally, intestinal AP isoenzymes are cleared quickly from the circulation. However, acute bowel diseases, especially when inflammatory in origin, can produce high serum AP levels. In this presented patient, the rise in serum AP levels preceded symptomatic manifestations of CMV colitis, and fell with successful therapy. Acute CMV disease in solid organ transplant recipients is common, can take many shapes, and needs to be diagnosed quickly. An unexplained rise in serum AP levels should lead to a search for inflammatory bowel disease, specifically CMV colitis, in transplanted patients.

Early Alteration in Bone Metabolism in Epileptic Children Receiving Carbamazepine Monotherapy Owing to the Induction of Hepatic Drug—Metabolizing Enzymes

The purpose of this study was to investigate, by a prospective, self-controlled method, whether early treatment with carbamazepine monotherapy can alter bone metabolism in ambulatory epileptic children with adequate sun exposure, based on the determination of total serum alkaline phosphatase and its bone isoenzyme activities. Serum total alkaline phosphatase and its bone, liver, and intestinal isoenzyme activities were evaluated in 22 epileptic ambulatory children (13 males and 9 females, aged from 5 to 12 years) before and at 3, 6, and 12 months of carbamazepine monotherapy. Serum concentrations of other biochemical markers of bone and liver metabolism, such as calcium, phosphorus, magnesium, γ-glutamyltransferase, alanine aminotransferase, aspartate aminotransferase, and lactate dehydrogenase, were also measured in the study participants before and at 6 and 12 months of treatment. Carbamazepine was prescribed at normal dosages (16.4—20 mg/kg/day). Serum total alkaline phosphatase activities were significantly increased at 3 ( P = .000), 6 ( P = .024), and 12 ( P = .037) months of treatment; serum bone alkaline phosphatase activities at 3 ( P = .000), 6 ( P = .008), and 12 ( P = .017) months of treatment; and serum liver alkaline phosphatase activities at 3 ( P = .000), 6 ( P = .049), and 12 ( P = .008) months of treatment, whereas serum intestinal alkaline phosphatase isoenzyme activity was significantly increased only at 3 months of treatment ( P = .035). Serum γ-glutamyltransferase activities were also significantly increased at 6 ( P = .000) and 12 ( P = .000) months of treatment. No significant changes in the concentrations of serum calcium, phosphorus, magnesium, alanine aminotransferase, aspartate aminotransferase, and lactate dehydrogenase were noted at 6 and 12 months of treatment. There was a significant correlation between serum γ-glutamyltransferase activities and serum total alkaline phosphatase activities ( r = .689, P = .000 at 6 months; r = .493, P = .020 at 12 months), bone alkaline phosphatase activities ( r = .700, P = .000 at 6 months; r = .466, P = .029 at 12 months), and liver alkaline phosphatase activities ( r = .427, P = .047 at 6 months; r = .425, P = .048 at 12 months). These findings indicate that ambulatory children who receive carbamazepine monotherapy, even when residing in a country with adequate sunlight, can have their bone metabolism altered early in the course of treatment, as indicated by the elevated activities of serum bone alkaline phosphatase isoenzyme. This early alteration in bone metabolism is probably due to the hepatic enzyme—inducing character of carbamazepine. ( J Child Neurol 2005;20:513—516).

Early Alteration in Bone Metabolism in Epileptic Children Receiving Carbamazepine Monotherapy Owing to the Induction of Hepatic Drug-Metabolizing Enzymes

The purpose of this study was to investigate, by a prospective, self-controlled method, whether early treatment with carbamazepine monotherapy can alter bone metabolism in ambulatory epileptic children with adequate sun exposure, based on the determination of total serum alkaline phosphatase and its bone isoenzyme activities. Serum total alkaline phosphatase and its bone, liver, and intestinal isoenzyme activities were evaluated in 22 epileptic ambulatory children (13 males and 9 females, aged from 5 to 12 years) before and at 3, 6, and 12 months of carbamazepine monotherapy. Serum concentrations of other biochemical markers of bone and liver metabolism, such as calcium, phosphorus, magnesium, gamma-glutamyltransferase, alanine aminotransferase, aspartate aminotransferase, and lactate dehydrogenase, were also measured in the study participants before and at 6 and 12 months of treatment. Carbamazepine was prescribed at normal dosages (16.4-20 mg/kg/day). Serum total alkaline phosphatase activities were significantly increased at 3 (P = .000), 6 (P = .024), and 12 (P = .037) months of treatment; serum bone alkaline phosphatase activities at 3 (P = .000), 6 (P = .008), and 12 (P = .017) months of treatment; and serum liver alkaline phosphatase activities at 3 (P = .000), 6 (P = .049), and 12 (P = .008) months of treatment, whereas serum intestinal alkaline phosphatase isoenzyme activity was significantly increased only at 3 months of treatment (P = .035). Serum gamma-glutamyltransferase activities were also significantly increased at 6 (P = .000) and 12 (P = .000) months of treatment. No significant changes in the concentrations of serum calcium, phosphorus, magnesium, alanine aminotransferase, aspartate aminotransferase, and lactate dehydrogenase were noted at 6 and 12 months of treatment. There was a significant correlation between serum gamma-glutamyltransferase activities and serum total alkaline phosphatase activities (r = .689, P = .000 at 6 months; r = .493, P = .020 at 12 months), bone alkaline phosphatase activities (r = .700, P = .000 at 6 months; r = .466, P = .029 at 12 months), and liver alkaline phosphatase activities (r = .427, P = .047 at 6 months; r = .425, P = .048 at 12 months). These findings indicate that ambulatory children who receive carbamazepine monotherapy, even when residing in a country with adequate sunlight, can have their bone metabolism altered early in the course of treatment, as indicated by the elevated activities of serum bone alkaline phosphatase isoenzyme. This early alteration in bone metabolism is probably due to the hepatic enzyme-inducing character of carbamazepine.

Alkaline phosphatase: placental and tissue-nonspecific isoenzymes hydrolyze phosphoethanolamine, inorganic pyrophosphate, and pyridoxal 5'-phosphate. Substrate accumulation in carriers of hypophosphatasia corrects during pregnancy.

Hypophosphatasia features selective deficiency of activity of the tissue-nonspecific (liver/bone/kidney) alkaline phosphatase (ALP) isoenzyme (TNSALP); placental and intestinal ALP isoenzyme (PALP and IALP, respectively) activity is not reduced. Three phosphocompounds (phosphoethanolamine [PEA], inorganic pyrophosphate [PPi], and pyridoxal 5'-phosphate [PLP]) accumulate endogenously and appear, therefore, to be natural substrates for TNSALP. Carriers for hypophosphatasia may have decreased serum ALP activity and elevated substrate levels. To test whether human PALP and TNSALP are physiologically active toward the same substrates, we studied PEA, PPi, and PLP levels during and after pregnancy in three women who are carriers for hypophosphatasia. Hypophosphatasemia corrected during the third trimester because of PALP in maternal blood. Blood or urine concentrations of PEA, PPi, and PLP diminished substantially during that time. After childbirth, maternal circulating levels of PALP decreased, and PEA, PPi, and PLP levels abruptly increased. In serum, unremarkable concentrations of IALP and low levels of TNSALP did not change during the study period. We conclude that PALP, like TNSALP, is physiologically active toward PEA, PPi, and PLP in humans. We speculate from molecular/crystallographic information, indicating significant similarity of structure of the substrate-binding site of ALPs throughout nature, that all ALP isoenzymes recognize these same three phosphocompound substrates.

Familial increased serum intestinal alkaline phosphatase: a new variant associated with Gilbert's syndrome.

Investigation of mild, inherited increased serum alkaline phosphatase activity partially combined with Gilbert's syndrome in one family showed, apart from a normal liver fraction, an intestinal isoenzyme pattern and an extra band in the agar electrophoresis. Analysis by agarose electrophoresis before and after incubation of neuraminidase showed that the extra fraction was an intestinal variant isoenzyme. The precise genetic background of the two disorders in this family could not be determined from the available data. Abnormal activities of (regular) intestinal alkaline phosphatase isoenzyme caused the increase in serum alkaline phosphatase in the absence of disease.

A monoclonal antibody against the surface of osteoblasts recognizes alkaline phosphatase isoenzymes in bone, liver, kidney, and intestine

Monoclonal antibodies against the surface of embryonic osteogenic cells have been used to characterize the osteoblastic lineage. One antibody, SB-1, reacts in frozen sections with a family of cells in bone, liver, kidney, and intestine which are identically stained by the histochemical substrate for alkaline phosphatase. In this report, biochemical and immunochemical evidence is presented to indicate that SB-1 is directed against an epitope on alkaline phosphatase which is shared by isoenzymes in a variety of chick tissues. In a solid-phase assay system, high dilutions (1:10 5) of ascites fluid were found to give significant binding of SB-1 to alkaline phosphatase extracted from chick limb or intestine. Partial purification of intestinal alkaline phosphatase on a Sepharose CL-6B column results in the co-elution of alkaline phosphatase enzyme activity and antibody-binding material; this indicates that SB-1 recognizes intestinal alkaline phosphatase rather than an impurity in the crude preparation. Furthermore, Western immunoblots of chick calvarial bone extract electrophoresed on a 5–20% SDS-polyacrylamide gel show that SB-1 reacts with a single 155 kD band which also is stained by the alkaline phosphatase histochemical substrate. In a similar set of experiments, SB-1 reacts with an intestinal alkaline phosphatase isoenzyme whose molecular weight is approximately 185 kD. From these studies, we conclude that SB-1 is specifically reactive with alkaline phosphatase isoenzymes present in bone, liver, kidney, cartilage, and intestine. The reactive epitope is stable to SDS denaturation, not associated with the active site of the enzyme, and dependent on disulfide bonds which impart secondary structure to the protein.

Intestinal alkaline phosphatase isoenzyme in patients with primary liver cancer during treatment with N10-propargyl 5, 8-dideazafolic acid (CB 3717).

Serum aspartate transaminase (AST), alkaline phosphatase (ALP), and gamma-glutamyl transferase (gamma GT) activities and the serum ALP isoenzyme pattern have been measured in eleven patients undergoing treatment with CB3717. There were increases in the activity of all three enzymes. The ALP isoenzyme pattern showed an increased contribution of the intestinal isoenzyme to the serum ALP activity. This may be due to increased intestinal mucosal leakage or impaired uptake and breakdown of the intestinal isoenzyme by the liver.

Prenatal diagnosis of cystic fibrosis by microvillar enzyme assay on a sequence of 258 pregnancies.

Prenatal diagnosis of cystic fibrosis by microvillar enzyme assay on amniotic fluid supernatant has been carried out on 258 sequential pregnancies with a 1 in 4 recurrence risk, all with known outcome. In general the three enzymes evaluated, gamma-glutamyltranspeptidase, aminopeptidase M and the intestinal isoenzyme of alkaline phosphatase, showed a high degree of concordance. However, there were two unusual patterns of microvillar enzyme activity; in seven cases a low gamma-glutamyltranspeptidase activity was associated with elevated values of intestinal alkaline phosphatase, and in ten cases there were isolated low values of intestinal alkaline phosphatase. The former pattern was found to be associated with cystic fibrosis in five cases, while the latter was associated with a normal outcome in all ten cases. A retrospective analysis of enzyme values suggested that the optimal system for minimizing false positives and false negatives was to define foetal cystic fibrosis as a sample where two of the three microvillar enzymes were below a cut-off of half the median value for the gestational week. If such scoring were applied to the cases where conventional microvillar enzyme patterns were observed, the false positive rate was 2.3% and the false negative rate 4.4% between 17 and 20 weeks of gestation.

Microvillar enzyme analysis in amniotic fluid and the prenatal diagnosis of cystic fibrosis.

The activities of two microvillar enzymes, gamma-glutamyltranspeptidase (GGTP) and alkaline phosphatase (ALP) have been determined in amniotic fluid (AF) samples from 39 pregnancies with a 1-in-4 risk of cystic fibrosis. Seventeen of these were investigated prospectively. A reduced proportion of the fetal specific intestinal ALP isoenzyme was found in 7 of a total of 13 pregnancies with cystic fibrosis and in one pregnancy of confirmed normal outcome. Eight of the affected pregnancies were tested for AF GGTP activity and depressed levels were found in 15. None of the 3 liveborn cystic fibrosis cases in the prospective series was identified by the ALP assay although 2 had significantly reduced GGTP activity. There were several amniotic fluid samples from cases of cystic fibrosis, trisomy 18 and normal outcome which had discordant GGTP and ALP results. Four of the 6 cases of cystic fibrosis misclassified by the ALP assay had amniocentesis at 15 or 16 weeks gestation. Evidence is presented which confirms a previous suggestion that amniocentesis after 17 weeks gestation improves the predictability of the ALP isoenzyme assay for the prenatal diagnosis of cystic fibrosis.

Prenatal diagnosis of cystic fibrosis using a monoclonal antibody specific for intestinal alkaline phosphatase.

A monoclonal antibody (AAP-1), specific for the intestinal isoenzyme of alkaline phosphatase (ALP), has been used to develop an immunoassay for amniotic fluid samples. Values in the immunoassay correlated closely with those obtained by direct determination of phenylalanine-inhibitable ALP. A panel of 124 control second-trimester amniotic fluids and 21 fluids with a 1 in 4 risk of a cystic fibrosis fetus were examined in the immunoassay. Eight of 10 affected cases had values below an arbitrary cut-off of one third median, while all the non-affected cases were above this level. Almost identical results were obtained by enzymatic determination of phenylalanine-inhibitable ALP. However, in both systems the false positive rate (control fluids with values below one third median), was unacceptably high. It is pointed out that at present the most effective system for the prenatal diagnosis of cystic fibrosis is achieved by measuring the ratio of intestinal to total ALP in amniotic fluid supernatants. This is probably best effected by enzymatic assay in the presence of phenylalanine and homoarginine inhibition.

Experimental ischaemia of the ileum and concentrations of the intestinal isoenzyme of alkaline phosphatase in plasma and peritoneal fluid.

Experimental ischaemia of the ileum and concentrations of the intestinal isoenzyme of alkaline phosphatase in plasma and peritoneal fluid.

Antigenic determinants of human placental and testicular placental-like alkaline phosphatases as mapped by monoclonal antibodies.

Placental alkaline phosphatase (PLAP) in humans shows a high degree of genetic polymorphism as disclosed by electrophoretic analysis. Human testes contain trace amounts of a PLAP-like enzyme, that although immunologically cross-reactive with PLAP, shows unique catalytic properties. As an alternative approach to study enzyme polymorphism we have developed monoclonal antibodies to purified allelic variants of PLAP. Five different monoclonal antibodies are described in this report. The antibodies react with different epitopes on the PLAP molecule. Both conformational dependent and independent determinants are detected. Two epitopes are modified when comparing the S and F allelic variants of PLAP. One epitope is common to PLAP and the intestinal isoenzyme of alkaline phosphatase. The five epitopes appear to be mapped on two rather distant antigenic domains. Combinations of any two antibodies binding to different domains give immunoprecipitates with PLAP on Ouchterlony tests and give good response in sandwich enzyme-linked immunosorbent assays. A study of PLAP-like enzyme in 32 individual testis samples indicates differences in four of the epitopes when compared with PLAP. Four types of testicular enzymes can be distinguished based on their reactivities. These results indicate structural differences between the testicular PLAP-like enzyme and PLAP. These differences are compatible with an underlying genetic mechanism.

Intestinal alkaline phosphatase and the abo blood group system — a new aspect

The influence of erythrocytes of different blood groups on the intestinal isoenzyme of alkaline phosphatase (EC 3.1.3.1) was investigated. Binding of the enzyme to erythrocyte membranes was demonstrated by sedimentation experiments, by enzyme electrophoresis, by gel chromatography and immunologically using a modified Coombs technique. We found that red cells of blood group A bind almost all intestinal alkaline phosphatase; erythrocytes of blood group B or O to a much lesser degree. This is in accordance with the fact that intestinal alkaline phosphatase is found more frequently in the serum of individuals of blood group O or B than in serum of persons of blood group A. We conclude that underlying the described blood group dependent differences in synthesis of intestinal alkaline phosphatase lies a mechanism involved in the regulation of the activity of intestinal alkaline phosphatase in human serum.

Studies of rat alkaline phosphatase. II. Some applications of the methods for detecting the isoenzymes of plasma alkaline phosphatase in rats.

This report describes the application of a test system which applies different substrates, inhibitors and acrylamide electrophoresis to the study of the alkaline phosphatase activity of rats. The system, which was specifically devised to characterise the alkaline phosphatase activity of small quantities of plasma from rats in toxicological studies, was also used to examine the plasma from rats under several physiological conditions, e.g. time of day, age, diet, etc. These physiological factors can influence the measurement of plasma alkaline phosphatase activity and should be taken into account when designing toxicological studies. Different experimental regimes were used to produce increases in liver, bone and intestinal alkaline phosphatase isoenzymes in the plasma of rats and these were reflected in the characterisations by the system.