Quantification of cell surface proteins with bispecific antibodies.

Flow cytometry is an established method for fast and accurate quantitation of cellular protein levels and requires fluorescently labeled antibodies as well as calibration standards. A critical step for quantitation remains the production of suitable detection antibodies with a precisely defined ratio of antigen-binding sites to fluorophores. Problems often arise as a consequence of inefficient and unspecific labeling which can influence antibody properties. In addition, the number of incorporated fluorophores necessitates a special normalization step for quantitation. To address these problems, we constructed different mono- and bivalent bispecific antibodies with binding site(s) for the cell surface antigens, cMET, EGFR1/HER1, ErbB2/HER2 or ErbB3/HER3 and with an additional digoxigenin-binding single-chain Fv fusion. The fluorophore Cy5 was covalently coupled to digoxigenin and quantitatively bound by the bispecific antibody. A panel of tumor cell lines was assessed under different culture conditions for absolute receptor expression levels of the indicated antigens and the data were set in relation to mRNA, gene count and immunoblot data. We could reproducibly quantify these receptors, omit the otherwise required normalization step and demonstrate the superiority of a 1 + 1 bispecific antibody. The same antibodies were also used to quantify the number of proteins in intracellular vesicles in confocal microscopy. The antibodies can be stored like regular antibodies and can be coupled with different digoxigenin-labeled fluorophores which makes them excellent tools for FACS and imaging-based experiments.

[Sneddon syndrome: vasculitis or thrombotic disorder?]. / Sneddon-Syndrom: Vaskulitis oder thrombotische Vaskulopathie?

BACKGROUND: Livedo reticularis generalisata (LR) in combination with affection of CNS is referred to as Sneddon's syndrome (SNS). Latest data suggest chronic progressive systemic disorder with occlusion of small and medium sized vessels (e.g., cutis, brain, kidneys, heart, eyes). No conclusive etiology is known, though there are correlations to the antiphospholipid syndrome, systemic secondary vasculitis and coagulopathies. Hereditary and toxic factors seem to play a role in pathogenesis in some cases. CASE REPORT: Diagnostic procedure and clinical course of a 56-year-old woman with dementia and hemiparesis proceeded by LR is reported. MRI-, SPECT- and TCD-findings were congruent with diffuse ischemic lesions of the brain due to affection of small- and medium-sized vessels. Histopathological specimens of the brain, meninges and cutis were non diagnostic. Some laboratory findings suggested vasculitis as an underlying cause. LR improved under immunosuppressive therapy with prednisolone and azathioprin. CONCLUSION: SNS does not seem to be a nosological entity. A differentiation between primary (idiopathic) and secondary SNS is useful for different therapeutical approaches.

[Results of surgical therapy of acute pancreatitis]. / Ergebnisse der operativen Therapie der akuten Pankreatitis.

Operative treatment of acute pancreatitis was carried out in 143 patients. The disease was caused by bilestones in 68%, by alcohol in 23%, idiopathic in 8%, and by different reasons (postoperative, helminthic disease) in 1% of the patients. The severity of the disease was classified into the grades I to III according to Kümmerle. The lowest rate of complications and lowest mortality rates were observed in operations performed within the first hours or after two weeks after the beginning of the disease. The alcoholic pancreatitis showed the lowest mortality rate (16%), despite of 81% of these patients suffering from the severe grades II and III. This may result from the decision in favour of early second look operations in these patients.

[Surgical treatment of bleeding stomach and duodenal ulcer--retrospective analysis of a five-year period]. / Die chirurgische Behandlung des blutenden Magen- und Duodenalulkus--Retrospektive Analyse über einen Fünfjahreszeitraum.

94 patients have been operated upon for haemorrhagic gastroduodenal ulcer disease in the Wenckebach-Krankenhaus during the years 1986-1990. In all but 6 patients the ulcer has been controlled by an emergency gastroscopy. 25 of these patients had to undergo emergency operation at once for persistent bleeding after gastroscopy. From the other patients, another 31 suffered rebleeding and had to be laparotomised in an emergency procedure, too. Only in 38 cases an interval-operation was possible. No one of these 38 patients died. The overall mortality-rate was 14.9% (14 patients). The mean age of our patients was 68 (26-90) years, none of the patients under 60 years died, whereas the mortality-rate of the elder was as high as 20%. The operative procedures were ulcer excision and suturing of the bleeding vessel and distal gastrectomy. There was no difference in mortality-rates between resective and non resective procedures. Our aim to operate upon the patients in an elective way could be achieved in about one third only. Many patients refused an operation after primary control of bleeding.

[Results of surgery of the hemorrhaging stomach and duodenal ulcer--a 10-year retrospective study]. / Ergebnisse der Chirurgie des blutenden Magen-und Gastroduodenalulkus--eine retrospektive Studie über 10 Jahre.

Operations were performed on 131 patients for haemorrhagic peptic ulcer at Wenckebach Hospital, between 1977 and 1986. Roughly two thirds of all patients had never had ulcer before. Persistent haemorrhage was recorded from about 20 per cent of these cases by emergency gastroscopy. Immediate surgical action had to be taken on 23.7 per cent. Interval interventions were possible for 28.2 per cent, while almost 50 per cent had to be laparotomised for early recurrent bleeding. Mortality in the wake of immediate emergency operations and surgery for recurrent bleeding was nearly twice as high as that in the context of interval interventions. Resection was performed on 70 per cent, haemostasis only on 26 per cent, and additional vagotomy on 14 per cent. Overall mortality amounted to 27.5 per cent including 36.6 per cent for women and 22.8 per cent for men. Ulcer localisation had no impact on mortality. The highest mortality rate, 38.2 per cent, was recorded from patients on whom only haemostasis had been performed, while 16.7 per cent were recorded from those to whom additional vagotomy had been applied. Lethality in the context of resection amounted to 25.6 per cent. Twenty-three patients had to be relaparotomised (17.6 per cent), which pushed up the mortality rate to 43.5 per cent.

Hepatocellular carcinoma and focal nodular hyperplasia associated with norethandrolone-therapy: a case report.

A 33-year-old woman was treated for severe aplastic anemia with norethandrolone over a period of four years, with a cumulative dose of 25 g. In the fifth year of therapy two intrahepatic tumors were detected and were classified as hepatocellular carcinoma and as focal nodular hyperplasia, respectively.

[Is there progress in the operative treatment of sigmoid diverticulitis]. / Gibt es Fortschritte bei der operativen Behandlung der Sigmadiverticulitis?

A high percentage of people in the civilized Western world develops colonic diverticula. Signs of illness are coming from inflammation around the diverticula. Resection therapy is indicated after several inflammation episodes. The operation has often to be performed in case of emergency: Ileus, bleeding, perforation. We recommend the primary resection of the sigmoid colon with an transverse colostomy or the Hartmann operation even in case of perforation or peritonitis. We operated 140 patients. The mortality rate in our hospital was 12% of the elective and 27% of the emergency operations.

Clinical experience with the tumor marker Ca 19-9 in an unselected group of more than 300 tumor patients.

Since March, 1983, in vitro tests for the quantitative determination of the tumor-associated antigen Ca 19-9 (carbohydrate antigen 19-9) were performed in more than 300 patients with confirmed tumors. These tests had as a goal determination of the diagnostic value of this new potential tumor marker Ca 19-9 with regard to specificity and sensitivity of malignant growths in the gastrointestinal tract, breast, and lung. The applicability of Ca 19-9 RIA in posttherapeutic case control for early registration of recurrences and metastases was examined. First analyses of our patient collective confirmed the assumption that increased serum concentrations of Ca 19-9 are detectable in gastrointestinal tumors as well as in mammary and bronchial carcinoma. Thus Ca 19-9 is a nonspecific tumor-associated antigen that by itself does not allow an organ correlation of malignancies of the gastrointestinal tract, breast, and lung.

[Anastomosis healing in the irradiated colon. III. Microangiographic studies. An animal experiment study]. / Untersuchungen über die Anastomosenheilung am bestrahlten Kolon. III. Mikroangiographische Untersuchungen. Eine tierexperimentelle Untersuchung.

During an experimentation on animals, the vascularization of anastomoses in the recto-sigmoidal region was investigated by microangiographic examinations after an irradiation corresponding to the therapeutic proceeding in man. The anastomoses were performed with and without fibrin coating one day, four weeks, eight weeks and four months following to irradiation. The examinations were performed on the third, seventh, 14th, and 21th postoperative day. The results of microangiography do not show any difference between irradiated and non-irradiated animals.

[Anastomosis healing in the irradiated colon. II. Influence of irradiation on new formation of connective tissue. An animal experiment study]. / Untersuchungen über die Anastomosenheilung am bestrahlten Kolon. II. Der Einfluss der Bestrahlung auf die Bindegewebsneubildung. Eine tierexperimentelle Untersuchung.

With regard to the permanent increase of perioperative irradiations in case of colorectal tumors, the possible influence of irradiation on the healing of anastomoses has been investigated. 230 rats were submitted to an irradiation of the rectosigmoidal region corresponding to the therapeutic method employed in human patients (focal dose of 50 Gy, in fractions of 4 x 2 Gy per week, interval of three weeks without irradiation). Rectosigmoidal anastomoses were performed one day, four weeks, eight weeks, and four months after the irradiation. The increase of connective tissue which is decisive for the stability of the anastomosis and which can be additionally induced by irradiation, was determined 3, 7, 14 and 21 days after the operation by identification of the hydroxyproline concentration within the anastomotic region. A comparison with nonirradiated animals shows that irradiation induces an additional formation of new connective tissue which becomes more intensive as the time interval since the finishing of irradiation is longer.

[Roentgenologic contribution to the diagnosis of gallstone ileus]. / Röntgenologischer Beitrag zur Diagnostik des Gallensteinileus.

Between 1969 and 1982 8 patients of ours were operated on for gallstone ileus. Signs found in the preoperative x-ray examination indicating this condition are analyzed retrospectively. The frequency and specificity of the x-ray signs found in the abdominal survey radiograph are compared with those reported in the literature. The value of oral contrast media administration in the detection of a cholecystointestinal fistula and localization of the obstruction is discussed.

[Study about the healing of anastomoses in the irradiated colon--experimentation on animals (author's transl)]. / Untersuchungen zur Anastomosenheilung am bestrahlten Kolon. Eine tierexperimentelle Studie.

Operations on preirradiated intestinal segments are followed by a high rate of complication. In order to determine the period after which an anastomosis on the clinically/therapeutically irradiated colon involves the greatest risk, 230 rats were submitted to a fractionated X-ray stationary-field irradiation of the colon with 50 Gy. The colon was anastomosed one day, four weeks, eight weeks, and four months respectively after the irradiation. A possible protective influence of fibrin on the healing of anastomoses was examined. The resistance to bursting pressure is taken as a criterion of the security of the anastomosis. It was determined on the third, 7th, 14th, and 21st day after the operation. The irradiation alone produces a markedly higher resistance to bursting pressure in the not operated control animals. After presurgical irradiation, the healing of anastomoses is delayed. There is no dependence on the period of time between irradiation and operation. From the 14th day after surgery, the resistance to bursting pressure is considerably higher than initially. In our experiment, the healing of anastomoses was not improved by an additional fibrin application.

[Primary tumors of the small intestine (author's transl)]. / Primäre Dünndarmtumoren.

From 1969 to 1979, 19 patients were operated for tumors of the small intestine: 8 were benign and 11 malignant. Of the benign, mesenchymal tumors predominated, the malignant tumors were epithelial. The difficulty of diagnostic confirmation and its effect of the prognosis of the disease are discussed with reference to the literature.

[Late radiation damage in the intestine. Clinical aspects, therapy and prognosis (author's transl)]. / Strahlenspätschaden am Darm. Klinik, Therapie und Prognose.

The causes, incidence, symptoms, treatment and prognosis of late radiation damage of the intestine are reported with reference to the literature. The possibilities of elimination are discussed. From 1969 to 1979, 48 of our own patients (34 women and 14 men) were operated for radiogenic late sequelae of the intestine. 27 of these patients required emergency laparotomy. The high mortality of around 42% is essentially due to fecal peritonitis already present at the time of operation.