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History, biochemical properties, techniques and normal range of carcino-embryonic proteins and their applications in clinical diagnosis are reviewed in detail. The importance of alpha1-fetoprotein, colon embryonic antigen, fetal sulfoglycoprotein, alpha2H-globulin and Regan-isoenzyme of alkaline phosphatase is discussed for carcinomas of the gastrointestinal tract, other malignant tumors and benign gastroenterological diseases. The results of prospective studies in cancer high risk patients and the indications for the determination of carcino-embryonic proteins in gastroenterology were described.

The Regan-Isoenzyme of alkaline phosphatase was determined immunologically measuring the antibody-fixed tumor alkaline phosphatase activity. In 68 blood donors the normal range varied from 0.1145 to 0.6351 (delta E/20 min at 405 nm, chi = 0.3748, standard deviation = 0.1013). 28.9% of 83 patients with different carcinomas showed a pathologically elevated Regan-Isoenzyme activity in the serum. The percentage of positive results as compared to previous reports is higher due to a more sensitive technique.

Using precipitating antibodies to human liver alkaline phosphatase (L-AP), a quantitative method is evaluated for the determination of L-AP-activity in feces. Normal controls (n = 42) exhibit a logarithmic normal distribution. Patients with intrahepatic cholestasis (n = 36) demonstrate an increase, patients with extrahepatic cholestasis (n = 23) a decrease of the fecal L-AP-activity. Variations of fecal L-AP-activities due to a varying water content of the stools could be excluded by simultaneous determination of intestinal AP-activity. Prae- and postoperative determinations in 12 patients with extrahepatic cholestasis yielded an increase of L-AP-activity after reconstructive surgery. In intrahepatic cholestasis, high fecal L-AP-activities are found together with relative low total AP-activities in the serum whereas in extrahepatic cholestasis low fecal L-AP-activities are associated with relative high total AP-activities in the serum.

An immunoprecipitation procedure is presented for the measurement of rat intestinal alkaline phosphatase (I-AP) in feces. Controls exhibit a logarithmic normal distribution of fecal I-AP. Single administration of an agent toxic for small intestinal mucosa like bleomycin or triparanol induces an increase of fecal I-AP on the 1st day and a marked reduction of I-AP activity on the following 3 days. After the 5th day, abnormal high fecal I-AP activities were observed during regeneration. In parallel, morphometry and measurements of I-AP activity in the homogenate of small intestinal mucosa were performed. Rat fecal I-AP excretion proved to be a simple, non-invasive, sensitive marker for toxic damage of the small intestinal mucosa.

A partially purified human fetal bone alkaline phosphatase preparation (B-APf) exhibits a class of antigenic determinants not found on any adult bone or liver isoenzymes as well as a set of determinants which it shares with adult bone and liver. The evidence for this is the quantitative immunotitration data which demonstrate the expected precipitation of L-AP, B-APa, B-APf and K-AP by the anti-B-APf reagent at equivalence. However, at high antibody excess, I-AP and P-AP are also precipitated. Conversely, by employing an immunosorbent column of Sepharose-purified L-AP, one can remove the component carrying the antigenic determinants shared with L-AP, B-APa and K-AP. Remaining in solution are antibodies which recognize determinant(s) shared by B-APf, P-AP and I-AP.

The intestinal alkaline phosphatase (I-AP) in feces was investigated in 16 patients with adult coeliac disease and 2 patients with gluten-sensitive enteropathy in dermatitis herpetiformis using an immunoprecipitation method. The mean concentration of I-AP activity in feces was reduced by 76% in patients with coeliac disease in comparison to normal controls (11.5 : 47.5 U/g). Patients with total villous atrophy (mean = 5.3 U/g) demonstrated a lower activity than patients with partial villous atrophy (mean = 14.3 U/g). Follow-up studies showed stable fecal I-AP-activities in symptom-free patients, whereas patients with relapses were characterized by fluctuating activities. The administration of a single oral dose of 31 g gluten is followed by a fecal I-AP-excretion pattern similar to toxic damage of the small bowel mucosa.

An immunoprecipitation method for the determination of human intestinal alkaline phosphatase (I-AP) in feces is described using anti-human I-AP IgG. Antibodies to human I-AP are monospecific after absorption with human placental alkaline phosphatase and show no cross-reactions with human liver alkaline phosphatase or alkaline phosphatase from E. coli. Normal controls demonstrate a logarithmic normal distribution of I-AP activity in feces. The standard deviation of the intra- as well as of the inter-assay variation is 11.7%. This method represents a simple quantitative non-invasive in vivo assay for brush border damage under experimental and/or clinical conditions.

Viral-hepatitis-like hepatic damage ("aprindin hepatitis") developed in a 50-year-old woman after taking aprindin, the cause of the jaundice being confirmed by the same signs occurring when the drug was taken again after initial withdrawal.

In comparison to normal controls (n = 71) the activity of intestinal alkaline phosphatase in feces is reduced in chronic bowel disease using an immunoprecipitation method: patients with Crohns disease (n = 40) or inactive ulcerative colitis (n = 29) demonstrate small changes of fecal intestinal alkaline phosphate activity in comparison to normal controls. Intestinal alkaline phosphatase is reduced in patients with active ulcerative colitis (n = 11) to 50%, in patients with uraemic enteropathy (n = 18) to 30% and in patients with coeliac disease (n = 14) to 20% of the activity observed in normal controls. During cytostatic treatment of malignant tumors, fecal intestinal alkaline phosphatase activity increases as a sign of toxic damage of the intestinal mucosa.

The isoenzymatic pattern of aldolase was determined by immunoprecipitation with specific anti-aldolase A, B and C sera in 21 pathological liver tissues and in the sera of normals (n equals 20), liver cirrhotics (n equals 52) and hepatoma patients (n equals 22). The increase of aldolase A in primary liver cell carcinoma is not reflected in the sera of these patients, cannot be used for diagnostic purposes and is not hepatoma-specific.

The structural relationship between human alkaline phosphatase isoenzymes from placenta, intestine, liver, bone and kidney was investigated by lectin-binding affinity chromatography. In addition, antibody-binding sites of the enzymes were studied using monospecific antisera against each of the isoenzymes. Evidence is offered for the existence of three classes of alkaline phosphatases: the placental isoenzyme, the intestinal isoenzyme and the liver-bone-kidney-type-isoenzyme: 1. A high affinity to bind to concanavalin A and lentil lectin characterizes the placental isoenzyme in contrast to the other isoenzymes. The intestinal isoenzyme remains totally unbound. The liver-bone-kidney-isoenzyme demonstrates a microheterogeneity with bound and unbound parts. A small unbound fraction can be detected in the placental isoenzyme, also when lentil lectin is used. 2. The placental isoenzyme and the isoenzymes purified from liver, bone and kidney are bound by wheat germ lectin-Sepharose, but not the intestinal isoenzyme. All isoenzymes are eluted as a homogeneous peak. 3. Using Helix pomatia lectin-Sepharose, all isoenzymes are unreactive except a minor fraction of kidney alkaline phosphatase. 4. Antibodies to the placental isoenzyme show a partial cross-reaction with the intestinal isoenzyme, but can be obtained monospecific after absorption. 5. Antibodies to the intestinal isoenzyme show a partial cross-reaction to the placental isoenzyme, respectively, but are monospecific again after absorption. 6. Antibodies to liver, bone or kidney isoenzymes show a complete cross-reaction, but are unreactive with the placental and intestinal isoenzyme; after absorption with a heterologous isoenzyme of this group, no further reaction can be demonstrated with any of the three isoenzymes. Thus, lectin-binding affinity identifies the same isoenzyme classes by their carbohydrate parts, as antibodies presumably do by the protein parts of the isoenzymes. Furthermore, lectin-binding affinity demonstrates a microheterogeneity of the placental isoenzyme with lentil lectin-Sepharose and of the liver-bone-kidney-type-isoenzyme with different lectins.

Carcinoembryonic antigen (CEA) levels were determined in 114 patients with confirmed lung cancer at the time of diagnosis using the CEA Ire-Sorin radioimmunoassay. Elevated CEA values were found in 47%. Most of the patients with high CEA levels had clinically detectable metastases. Ferritin was detectable by the Laurell-electrophoresis in the serum of 58 out of 81 (72%) of the patients with confirmed lung cancer at the time of diagnosis. Ferritin levels were significantly higher in patients with metastases. Serial measurements of CEA and ferritin during radio- and chemotherapy showed that the assay may be useful to evaluate the effects of therapy. Because of some false negative results both CEA and ferritin determinations should be used only in context with other clinical and laboratory findings.

A new and simple method for the purification of human liver alkaline phosphatase using butanol extraction, acetone and ammonium sulfate precipitations, anion exchange chromatography and gel filtration is described.

Levels of carcinoembryonic antigen (CEA), alpha-fetoprotein (AFP), ferritin and alpha 2-pregency associated glycoprotein (alpha-2-PAG) were determined in patients with confirmed lung cancer at the time of diagnosis and in serial determinations during and after radio- or chemotherapy. Whereas AFP levels were not elevated in patients with lung cancer, increased levels of CEA, ferritin and alpha-2-PAG were found in more than 50% of the patients. The results suggest that determination of CEA, ferritin and alpha-2-PAG in the serum of patients with lung cancer may be useful to detect metastases or recurrences and to monitor the results of treatment. Furthermore, in this study CEA and ferritin could be demonstrated in extracts of lung tumor tissues by specific antisera.

Crystalline human alkaline phosphatase from placenta and intestine was isolated by butanol extraction, acetone precipitation, a heat step (for the placental enzyme), ammonium sulfate precipitation, anion exchange chromatography, gel filtration and crystallisation with ammonium sulfate. Rabbit antibodies showed a partial cross-reaction between both enzymes in double diffusion, quantitative precipitation experiments and in serial precipitin curves. There was no reaction with human alkaline phosphatases from liver, kidney and bone. Three phenotypes of placental alkaline phosphatase with different electrophoretical mobility exhibited identical immunological reactions with their respective antisera. Monospecific antisera were obtained by absorption with crystalline intestinal or placental alkaline phosphatase. These monospecific antisera against the placental or the intestinal alkaline phosphatase can be used for an immunological determination of these two alkaline phosphatases without contamination by other alkaline phosphatases in human serum.

The preparation of monospecific antisera for immunoassays of human placental (P-AP, Regan) and intestinal (I-AP) isoenzymes necessitates complete absorption of the antisera with heterologous antigen. Large scale preparation of pure human I-AP for absorption of anti-P-AP sera is not practical since a large amount of fresh intestine is required. Calf I-AP cross-reacts with human I-AP, but evidence is presented that this antigen is not suitable for absorption of anti-human-P-AP sera. In contrast, human I-AP can be extracted from faeces of patients under polychemotherapeutic treatment and used for the complete absorption of anti-human-P-AP sera. Monospecific anti-human-I-AP sera, suitable for immunofluorescence studies, can be obtained by a two-step affinity chromatography procedure: (1) human I-AP is removed from an extract of small intestine by antibodies to cross-reacting calf-I-AP, and (2) this small intestinal mucosal extract, free of human I-AP, can be used for the absorption of anti-human-I-AP sera. Finally, we present evidence that antibodies to calf-I-AP precipitate human-I-AP and can be used in immunoassays for I-AP in man.

(1) Passive hemagglutination and radioimmunoassay are suitable methods for the detection of AFP in the low concentration range. (2) In 3.72% of the cases a clinically unknown carcinoma was found in an unselected group of patients with liver cirrhosis. (3) 21.9% of the patients showed AFP elevations up to 2000 ng/ml. In 10.6% of this group, increasing titers demonstrated a primary liver cell carcinoma. In 89.4% a transitory rise of AFP was not associated with tumor growth. Levels return to normal values within three months in 90% of the cases. (4) Transitory AFP elevations are not correlated to clinical conditions (praecoma, coma, delirium, bleeding, ascites, shunt) or to biochemical parameters (GOT, GPT, bilirubin, prothrombin complex time, gamma-globulin). (5) A temporary rise in AFP is more frequently observed in groups with high hepatoma incidence than in groups with low hepatoma incidence. (6) Therefore, it may be suggested that a transitory rise of AFP could reflect a "primary reaction" of carcinogenesis. (7) Primary liver cell carcinoma is found to be more frequent in posthepatitic than in postalcoholic, cryptogenic, and other cirrhosis and to be more frequent in australia-antigen positive than in australia-antigen negative cases. (8) Routine serological tumor antigen screening of patients with a precancerous disease is useful.

In 38 children and adolescents with cystic fibrosis, in 19 normal children and 62 healthy adults the serum alpha1-fetoprotein concentrations were measured by radioimmunoassay. In cystic fibrosis patients 97.5% and in normal children 95% of the alpha1-fetoprotein values were within the normal range for healthy adults (1--9 ng/ml). Critical judgement of the reported findings in literature and our own results demonstrate that the investigation of alpha1-fetoprotein in the serum cannot serve for detecting homozygotes of cystic fibrosis genes or heterozygote carriers.

The incidence of primary liver cell carcinoma was investigated in a prospective study over 6 yr and 5 mo in 403 clinically unselected patients derived from a homogeneous population by means of serial determination of alpha 1-fetoprotein (AFP) by radioimmunoassay. The diagnosis of liver cirrhosis was proved in 90% by laparoscopy and/or histology and/or autopsy. The incidence of primary liver cell carcinoma in liver cirrhosis in the clinically studied patients was 4.47%, significantly lower than in the autopsy material (11.03%; p less than or equal to 0.025). In the follow-up study, all patients with increasing AFP concentrations exhibited a primary liver cell carcinoma. A transitory rise of AFP (higher than 50 ng/ml) was observed in 15.1% of patients with liver cirrhosis without primary liver cell cancer. In contrast to the results of animal experiments, this transitory rise of AFP was not followed by malignant transformation of the cirrhotic tissue. Posthepatitic liver cirrhosis was observed in 21.57%, postalcoholic liver cirrhosis in 42.93%, and cryptogenic liver cirrhosis in 27.30%. Liver cirrhosis of other etiology occurred in 8.19%. The incidences of primary liver cell cancer in these 4 groups were 4.94, 4.62, 5.45, and 0%, respectively. These differences are not statistically significant, although in absolute figures postalcoholic liver cirrhosis is the main cause of primary liver cell carcinoma in this sample from West Germany. HBs antigen-positive liver cirrhosis was more often associated with primary liver cell cancer than HBs antigen-negative liver cirrhosis (6.58 versus 3.96%); this difference also is not statistically significant. Observations of larger groups of patients may show a higher risk of developing primary liver cell carcinoma in those with a combination of alcohol abuse and HBs antigenemia and/or acute hepatitis in the history. Patients without these 2 risk factors had an incidence of primary liver cell carcinoma of 2.61%; those with 1 risk factor, 5.77%; and those with both risk factors, 10.71%.

Single administration of ethanol or alcoholic beverages (5g ethanol/kg body weight) induces a decrease of the 24h excretion of fecal intestinal alkaline phosphatase (I-AP) activity of 28% (P less than 0.05) in comparison to controls (0.9% saline). Administration of higher amounts of 20% (v/v) ethanol (8g/kg body weight on 3 consecutive days) yields a decrease of fecal I-AP excretion up to 82% (P less than 0.005) in comparison to controls (saturated glucose solution). The interpretation of these results as a toxic effect of ethanol to small intestinal mucosa was supported by measurement of enzymatic activity in the small intestinal mucosa and by morphometric data.

Immunoreactive serum levels of human chorionic gonadotrophin (HCG), its alpha- and beta- subunits (alpha-HCG and beta-HCG), calcitonin (CT), parathyroid hormone (PTH), prolactin (PRL), adrenocorticotropic hormone (ACTH), and growth hormone (GH) were increased in 8 to 68% of 44 patients with hepatocellular carcinoma. With the exception of two patients, ACTH and PRL levels were only moderately increased, while alpha-HCG, GH, ACTH and PRL levels were not significantly different from the levels found in cirrhosis suggesting that metabolic effects due to impaired liver function may be responsible for their increase in liver cirrhosis and primary liver cell carcinoma. In contrast, HCG, beta-HCG, CT and PTH were associated with a higher incidence of elevated immunoreactive hormone levels than the other peptide hormones; higher concentrations were noted in tumor patients than with liver cirrhosis alone. Therefore, we suggest that metabolic effects due to cirrhosis may influence the serum levels and be more important than ectopic secretion by hepatocellular carcinoma.

Carcinoembryonic antigen (CEA) was determined at the time of diagnosis in 90 patients with lung cancer. CEA levels were elevated in 47 (52%). High CEA levels were often associated with the presence of metastases. Serial CEA determinations in 40 patients, before and after radiotherapy and chemotherapy, revealed that patients who responded to treatment had a decreased CEA level, whereas those with progressive tumour growth had increased ones. CEA assay may, therefore, be useful in detecting metastases and in helping to assess the effect of treatment in patients with lung cancer.

Four different immunological methods for the determination of the placental isoenzyme of alkaline phosphatase (Regan isoenzyme) were compared in 64 normal blood donors, 23 healthy laboratory and medical staff workers and 68 pregnant women: a. Inhibition by soluble antibodies to the placental enzyme. b. Precipitation with soluble antibodies. c. Precipitation with immobilized antibodies. d. Measurement of the activity of the placental alkaline phosphatase following binding to an immunoabsorbent that has been obtained by polymerization of anti-placental-alkaline phosphatase gamma-globulin using glutaraldehyde. The immunoabsorbent method yielded the best results. The optimal conditions were evaluated for the measurement of the activity of immunoabsorbent-fixed placental (tumoral) alkaline phosphatase activity. This method was applied to 209 normal blood donors and to 239 patients with different malignant tumors: 25.5 percent of the cancer patients exhibited an elevated Regan -isoenzyme activity in the serum.