Europ. J. Pedia$. 122, 241--248 (1976) 9 by Springer-Verlag 1976

Serum Alphal-Fetoprotein in Cystic Fibrosis G. KnSpfle, H. W. Rotthauwe, and F.-G. Lehmann Department of Pediatrics, University of Bonn Department of Medicine, University of Marburg Received February 2, 1976 Abstract. In 38 children and adolescents with cystic fibrosis, in 19 normal children and 62 healthy adults the serum al-fetoprotein concentrations were measured by radioimmunoassay. In cystic fibrosis patients 97.5~o and in normal children 95~o of the al-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 al-fetoprotein in the serum cannot serve for detecting homozygotes of cystic fibrosis genes or heterozygote carriers. Key words: Alphal-fetoprotein - - Cystic fibrosis. Zusammen/assung. Bei 38 Kindern und Jugendlichen mi~ cystischer Fibrose, 19 gesunden Kindern und 62 gesunden Erwachsenen wurden die Serumkonzentrationen des Alphal-Fetoproteins im Radioimmunoassay bestimmt. 97,5~o der Alphal-Fetoprotein-Werte der Patienten mit cystischer Fibrose und 95~o der MeSwerte der gesunden Kinder lagen innerhalb des Normbereiches (1--9 ng/ml), der fiir gestmde Erwachsene ermittelt worden war. Aufgrund unserer eigenen Untersuchungsergebnisse und bei kritischer Bewertung der bisher publizierten Befunde erscheint die Alphal-Fetoprotein-Bestimmung im Serum zur Erfassung der homozygoten und heterozygoten Gentr~ger der cystischen Fibrose nieht geeignet. Introduction

High levels of al-fetoprotein (AFP), a carcino-embryonic protein have been found in fetal serum after 4 weeks of gestation, in the serum of newborns, and pregnant women [1--4]. Fetal hepatocytes are the major site of embryonal AFP-synthesis. During the embryonic period in humans the highest A F P concentrations in fetal serum (3 mg/ml) occur after about 1 2 - - i 6 weeks of gestation [1, 5]. I n the older fetus the level of A F P decreases and in infants at term the concentration ranges between 20 and 170 ~g/ml [1--3]. Using immunoprecipitation techniques [limit of sensitivity in double diffusion (micro-Ouchterloney technique): 7500 ng/ml, limit of sensitivity in electroimmuno-osmophoresis: 2000 ng/ml] A F P cannot be detected in the serum of normal infants after 3 weeks of life. The determination of A F P b y more sensitive immunologic techniques such as the radioimmunoassay has shown the presence of A F P concentration in sera of normal adults and children. The upper limit of the A F P concentration in normal h u m a n serum is 10 and 20 ng/ml, respectively [6--9]. I n normal pregnancies between 10 and 14 weeks of gestation the A F P level in maternal serum increases and the highest A F P concentrations (up to 500 ng/ml) have been found in the 25th to 35th week [4, 6, 10]. I n children and nonpregnant individuals serum A F P levels above 2 ~g/ml are virtually c h a r a c -

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teristic for primary liver cell carcinoma and teratoblastoma containing yolk sac tissue [11--15]. I n patients with other types of carcinomas, A F P values above 2 ng/ml are rare (in stomach carcinomas about 3.5--4%, in gallbladder and bile duct carcinomas about 3% and in other carcinomas with or without liver metastases less t h a n 0.3%). A transient rise of A F P concentrations up to 100 ng/ml, in rare cases up to 500 or 2000 rig/m1 was seen in about 15--25% of the sera in patients with benign liver diseases such as viral hepatitis, chronic active liver disease, alcoholic hepatitis or liver cirrhosis [16--19]. Raised serum A F P levels (40--290 ~g/ml) after 3 weeks of life were detected in infants with neonatal hepatitis, too, but not in infants with biliary atresia [20, 21]. Moreover, increased levels of A F P were found in the sera of children with ataxiatelangiectasia and hereditary tyrosinemia [22--24]. I n pregnancy disorders maternal A F P levels m a y increase and exceed the 97.5 percentile of A F P concentrations appearing in normal pregnancy. I n imminent spontaneous abortions, in diabetic and Rhimmunized pregnancies, and especially in intrauterine fetal death the maternal A F P levels m a y be raised up to 9000 ng/ml [4, 6, 10]. Abnormally high amniotic fluid A F P values are associated with various types of fetal anomalies such as neural tube malformations (anencephaly, ope n spina bifida, myelomeningocele), 45,X0 chromosome constitution, congenital nephrosis of Finnish type, esophageal atresia, duodenal atresia, fetomaterual transfusion, intrauterine fetal death, and severe Rh-immunization [25--31]. Recently, Chandra et al. [32] reported significantly increased levels of A F P in the serum for both homozygote and heterozygote carriers of cystic fibrosis (CF) genes in Newfoundiand, Canada. These findings were confirmed b y Smith [33] in the USA, whereas authors in England, Scotland, and also in Canada and in the USA, were unable to detect an increased serum A F P concentration in any of their CF patients or their relatives [34--37]. Since so far no investigations in CF patients in central Europe have been performed, we have measured the A F P levels in children and adolescents with CF in the western p a r t of the Federal Republic of Germany. The values were compared with those in healthy adults and normal children. I n a " L e t t e r to the Editor" in the British Medical Journal the different results in Canadian and North American CF patients and patients with CF in central and northwestern Europe are to be found [38]. Material We examined the sera of 38 CF patients with the disease (16 g~, 22 ~) in various stages of severity (mean age 4- SD: 9.06 :~ 3.76 years). Except for one Turkish boy, all patients with CF were of German origin. The diagnosis of CF was based on both clinical symptoms of pulmonary and pancreas involvement and sweat sodium chloride levels above 80 mmol/1 on pilocarpine iontophoresis. Pancreatic enzyme replacement therapy and lipasoluble vitamin supplements were necessary in all CF patients except for 3 children. An oral medication of bronchiolytic agents (Bisolvon | was given to all patients with CF. Thirteen children were on continous antibiotic therapy. In addition to daily postural drainage of bronchial secretions all C~ patients had to undergo aerosol inhalations with N-acetyl cysteine (~istabronco | Mucolytieum Lappe| Sixteen patients were on nocturnal ultrasonic mist tent therapy. Percutaneous liver biopsy was performed in 9 CF patients 1. In 2 other patients aged i The histologic investigations were performed by Prof. F6disch, Department of Pediatric Pathology, University of Bonn.

Serum Alphal-Fetoprotein in Cystic Fibrosis

243

5 and 7 years an increase of liver enzymes in the serum was observed over several years as a sign of liver damage (GOT 30--50 mU/ml, Gamma-GT 50--100 mU/ml). Serum total protein and albumin concentration were normal in all CF patients; increased serum levels of gamma globulin and al-antitrypsin were found in the majority of CF patients due to chronic bronchitis, peribronchial inflammations, and recurrent pneumonia. Furthermore, we examined the serum AFP levels in a group of 10 female and 9 male children aged 2--12 years (mean age :[: SD: 8.35 :~ 3.64 years). All subjects were recovered inpatients at the Department of Pediatrics, University of Bonn. The clinical examination revealed normal conditions and all children were discharged 1 or 2 days later. Their personal history did not give any clue of recently acute or chronic liver diseases or metabolic disorders. At least 1 week before blood samples were taken, any medication had been stopped. Usually the venous blood samples were taken in the morning from fasting subjects. Serum samples were kept deep frozen until analysis. The normal adults were of 62 healthy blood donors.

Methods The quantitative determination of AFP in the serum was carried out by radioimmunoassay with lsSJ-labeled AFP~ in the double antibody technique. The first antibody was a rabbit antihuman AFP-serum~, the second antibody a goat antirabbit-y-globulin2. The test was carried out in 0.1 M borate buffer (pH 8.6) containing 0.5% beef albumin as described by Nishi and Hirai [39] and Ishii [40]. Incubation periods were always 24 h for the first antibody, the radioactive labeled AFP, and the second antibody. Crystalline AFP, isolated according to Lehmann [41, 42], was used as standard. The concentration of crystalline AFP (local standard) was 90 t~g/ml by Mancini's radial immnnodiffusion technique [43] and 82 ~g/ml by radioimmunoassay in the International Standard Preparation 72/225 of the World Health Organisation [44]. In addition, the sera of all 38 patients with CF were examined both by eleetroimmuno-osmophoresis(limit of sensitivity2000 ng/mI) and by double diffusion (limit of sensitivity 7500 ng/ml) with rabbit antihuman AFP-sernm as described by Lehmann and Lehmann [45]. Serum AFP values in our study groups were not normally distributed. Therefore we used for statistical analysis the nonparametric U-test of Wilcoxon et al. [46], which tests the zero hypothesis whether two independent samples come from the same entity at a given significance level. Out of three independent groups for which two multiple tests {healthy adults - - CF patients; normal children - - CF patients) had to be done and for every single test a significance level of 2.5% was chosen, to make sure that the overall error in ease of significance does not exceed the overall level of 5%.

Results F i g u r e 1 shows b y h i s t o g r a m the serum A F P c o n c e n t r a t i o n s i n h e a l t h y adults, n o r m a l children, a n d p a t i e n t s with CF expressed as relative s u m frequency. I n CF p a t i e n t s 97.5% a n d in n o r m a l children 95% of the A F P values were w i t h i n t h e n o r m a l range for h e a l t h y a d u l t s ( 1 - - 9 ng/ml). T a b l e 1 presents m e a n values a n d s t a n d a r d deviations, m e d i a n s , a n d ranges of t h e serum A F P c o n c e n t r a t i o n s i n our three groups. U s i n g the U - t e s t of Wileoxon et al. t h e statistical analysis revealed significantly lower A F P c o n c e n t r a t i o n s in the s e r u m of CF p a t i e n t s b o t h i n c o m p a r i s o n to the values of n o r m a l children a n d to those of h e a l t h y adults. I n our series i n 7 of 9 CF p a t i e n t s t h e histologic lesions o b t a i n e d b y liver biopsy were u s u a l l y mild. M a i n l y there was a f a t t y i n f i l t r a t i o n of liver cells (Table 2). The serum A F P c o n c e n t r a t i o n s i n those p a t i e n t s (n = 7) h a v i n g histologie liver changes ( m e a n v a l u e =t= SD: 3 ~ 2 ng/ml, m e d i a n 2 ng/ml, r a n g e 1 - - 6 ng/ml) were n o t significantly different from those i n o t h e r C F p a t i e n t s ( P > 0.05). Con2 Dianabot-Laboratories, Tokyo, Japan.

244

G. Kn5pfle et al. 9 sum frequency

I~1 I"1

CF patients normal children normal adults

1.0 0.9 0.8

0.7 0.6 0.5

0.3 0.2 0.1

1t 1

2.

3

4

5

.6

7

8

9

10

AFP ng/ml

Fig. 1. Serum AFP concentrations in healthy adults (n : 62), normal children (n = 19), and patients with CF (n = 38) expressed as relative sum frequency by histogram

cerning age d i s t r i b u t i o n there was n o significant difference b e t w e e n the p a t i e n t s with CF a n d t h e n o r m a l children ( P > 0.05). T h e sera of all 38 p a t i e n t s with CF e x a m i n e d a d d i t i o n a l l y b y eleetroimmuno-osmophoresis a n d b y double diffusion were n e g a t i v e i n b o t h techniques.

Table 1. Serum cq-fetoprotein concentrations in healthy adults, normal children, and patients with cystic fibrosis Group

Number

A_FP (ng/ml)

Healthy adults

62

Mean value =t= SD Median Range

4.52 • 1.92 4.0 1.0 - - 9.0

Patients with cystic fibrosis

38

Mean value :]: SD Median Range

3.08 :~ 2.27 3.0 1.0 - - 10.0

Normal children

19

Mean value :J= SD Median Range

6.10 =[= 1.96 6.0 3.0 - - 10.0

Probability of zero hypothesis (U-test of Wilcoxon et al.)

P < 0.025

P < 0.025

Serum Alphal-Fetoprotein in Cystic Fibrosis

245

Table 2. Serum arfetoprotein concentrations and histologic findings obtained by liver biopsy in 9 patients with cystic fibrosis Patient

Age (years)

AFP (ng/ml)

ttistologie findings

E.S. O.R. R.S. 1~. I-I. B.A. O.M. L.M. B.J.

69/12 159/12 132/12 117/12 79/19 54/1~ 64/12 67/19

5 1 2 5 4 1 2 6

79/19

1

normal normal mild fatty infiltration moderate fatty infiltration and portal fibrosis mild fatty infiltration moderate fatty infiltration severe fatty infiltration concretions plugging small bile ducts and mild portal inflammatoI7 changes unspecific mild portal inflammatory changes

g.F.

Discussion

Cystic fibrosis or mucoviscidosis is the most frequent autosomal recessive inherited disease in Caucasian populations. Clinical manifestations of CF occur only in homozygous gene state, whereas heterozygote carriers of CF genes are asymptomatic. The gene frequency in Caucasians is approximately 2 - - 5 % . The incidence of CF is estimated about 1 in 2000 live births. The basic biochemical defect is still unknown. In homozygotes there is a generalized disorder in the exocrine system, in which the secretory fluids have abnormal compositions. Sweat, lachrimal, and salivary glands produce secretions of increased electrolyte content and mucus glands secrete a very tenacious mucus. The abnormally viscous secretions may obstruct the gland ducts in a progressive cystic dilation and may result in fibrosis of gland parenchyma. In the majority of CF patients postmortem examinations revealed histologic changes in the liver [47 19]. Hepatic abnormalities include distension of intrahepatie bile ducts, unspecific f a t t y infiltration, focal biliary fibrosis, and multilobular cirrhosis with concretions, the typical liver lesion in CF [48, 50---52]. In our material the serum AFP concentrations in CF patients were slightly, but significantly lower than those in healthy adults and normal children. This finding m a y be explained by the fact, t h a t in contrast to the two normal groups the quantitative determination of serum A F P levels in CF patients was carried out by using the same crystalline standard, but in another test series with a different antiserum and a different radioactive labeled AFP. Our findings correspond with those of several other authors [34--37]. These authors also, in contrast to both Chandra et al. [32] and Smith [33] have been unable to detect by radioimmunoassay and enzyme immunoassay an increased serum concentration of A F P in any of the 130 CF patients tested in England, Scotland, Canada, and in the USA [34---37]. The serum AFP levels of their patients with CF, too, were within the normal range for healthy controls. At the moment there is no completely satisfactory explanation for the discrepancy in the results as Chandra et al. [32] in Canadian patients with OF have apparently used the same rabbit antiserum and standards as Brock et al. [35] in CF patients in Scotland. Various grades of severity of the disease

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or genetic differences cannot explain the different results concerning the serum A F P levels in patients with CF. The m a j o r i t y of the ancestors of the CF patients who have been tested b y Smith in the U S A [33] and b y Chandra et al. [32] in Canada m a y huve their origin in central E u r o p e and the northwestern p a r t of E u r o p e just those of the other CF patients tested in the same geographical areas [37]. B@langer et al. [37] discuss the possibility t h a t the antisera used b y C h a n d r a et al. [32] and b y Smith [33] react t o w a r d an antigen other t h a n A F P which m a y occur in sera of CF patients and which m a y be a m a r k e r of the disease. Critical judgement of the reported findings in literature and our own results demonstrates t h a t the investigation of al-fetoprotein in the serum cannot serve for detecting homozygotes of CF genes or heterozygote carriers.

Acknowledgements. The authors acknowledge the technical assistance of Mrs. A. Odenthal and l~rs. M. Bensch.

References 1. Gitlin, D., Boesman, IV[.: Serum AFP, albumin and 7G-globulin in human conceptus. J. clin. Invest. 45, 1826 (1966) 2. Adinolfi, 1K.,Gardner, B. : Studies on the properties of human fetal serum protein. Develop. reed. Child. Iqeurol. 9, 609 (1967) 3. Marklein, G., Rings, M.: Quantitative immunologische Bestimmung des Alphal-Feteproteins im Serum reifer und unreifer Iqeugeborener sowie im Serum yon S~uglingen und Kindern. Z. Kinderheilk. 113, 327 (1972) 4. Sepp~il~, M., 1%uoslahti, E.: Alpha-fetoprotein in normal and pregnancy sera. Lancet 1972 I, 375 5. van Furth, 1%, Adinolfi, 1K.: In vitro synthesis of fetal alpha-l-globulin in man. Nature (Lend.) 222, 1296 (1969) 6. Purves, L. 1%, 1)urves, M.: Serum alpha-fetoprotein. VI. The radioimmunoassay. Evidence for the presence of AFP in the serum of normal people and during pregnancy. S. Aft. reed. J. 46, 1290 (1972) 7. 1%uoslahti, E., Seppiil{i, IV[.: Studies on careino-fetal proteins. III. Development of a radioimmunoassay for AFP. Demonstration of al-fetoprotein in serum of healthy human adults. Int. J. Cancer 8, 374 (1971) 8. 1%uoslahti, E., SeppEl~, M.: AFP-detection in normal serum. Nature (Lend.) 235, 161 (1972) 9. l~asseyeff, 1%., Gilli, G., Krebs, B., Boner, C., Zrihen, It. : Evolution en fonction de l'age du taux s6rique physiologique de l'alpha-foetoprot~ine chez l'homme et le rat. In: Alphafeteprotein (ed. 1%.Yfasseyeff), pp. 313--322. Nice: Cell de l'INE1%~ 10. SeppElE, l~., 1%uoslahti, E.: 1%adioimmunoassay of maternal serum alpha-fetoprotein during pregnancy and delivery. Amer. J. Obstet. Gynee. 112, 208 (1972) 11. Abelev, G. 1. : Production of embryonal serum a-globulin by hepatomas: Review of experimental and clinical data. Cancer 1%es. 28, 1344 (1968) 12. Smith, J.B., O'Neill, 1%.T.: Alpha-fetoprotein. Occurrence in germinal cell and liver malignancies. Amer. J. Med. ~1, 767 (1971) 13. Ruoslahti, E., Sepp~lii, !K., Vuopio, P., Saksela, E., Peltekallio, P.: 1%adioimmunoassay of alpha-fetoprotein in primary and secondary cancer of the liver malignancies. J. nat. Cancer Inst. 49, 623 (1972) 14. Gassner, 1~., Grob, P . J . : al-fetoprotein, hepatom and teratom. Schweiz. reed. Wschr. 102, 465 (1972) 15. Lehmann, F.-G. : Kristallisation yon al-Foetoprotein aus dem Plasma eines ~'atienten mit prim~rem Leberzellkarzinom. Klin. Wsehr. 49, 609 (1971) 16. 1%uoslahti, E., Sepp~l~, M., R~sS,nen, J. A., Vuopio, P., I-Ielske, T.: Alpha-fetoprotein and hepatitis B antigen in acute hepatitis and primary cancer of the liver. Scand. J. Gastroent. 8, 197 (1973b)

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17. Geffroy, Y., Denis, P., Colin, R., Sauger, F., 1VIatray, F., Fondimare, A.: Prdsence d'alpha1-fotoprotdine che~ l'adulte au cours d'une h~patite virale traitde par corticothdrapie. Presse todd. 78, 1107 (1970) 18. Masopust, J., Radl, J., Houstek, J. : Occurrence ofal-foetoprotein in some infants suffering from hepatopathy. Prot. Biol. fluids 18, 239 (1971 a) 19. Lehmann, F.-G.: Der Naehweis von al-Foetoprotein bei internen Erkrankungen aktueller Stand des Problems. Internist (Berl.) 13, 332 (1972) 20. Kohn, J., Weaver, P.C.: Serum alpha-fetoprotein in hepatocelIular carcinoma. Lancet 1974 II' 334 21. Zelter, P. M., Neerhout, t~. C., Fonkalsrud, E. W., Stiehm, E. R. : Differentiation between neonatal hepatitis and biliary atresia by measuring serum alpha-fetoprotein. Lancet 1974 I' 373 22. Bdlanger, L.: Tyrosin~mie h~r~ditaire et alpha-foetoprot~imie. Path. et Biol. 21, 457 (1973) 23. Bdlanger, L., B~langer, M., Prove, L., Larochelle, J., Tremblay, M., Aubin, G. : Tyrosindmie hdr~ditaire et alpha-l-foetoprot6ine. Path. et Biol. 21, 449 (1973) 24. Waldmann, T.A., McIntire, K. 1~.: Serum alpha-foetoprotein levels in patients with ataxia-telangieetasia. Lancet 1972 II' 1112 25. Seller, M. J., Creasy, M. R., Alberman, E. D. : al-fetoprotein levels in amniotic fluids from spontaneous abortions. Brit. med. J. 1974 II, 524 26. Brock, D. J. tt., Suteliffe, R . G . : Alpha-fetoprotein in the antenatal diagnosis of anencephaly and spina bifida. Lancet 1972 II, 197 27. Allan, L. D., Ferguson-Smith, M. A., Donald, J., Sweet, E. M., Gibson, A. A. M. : Amniotiefluid alpha-fetoprotein in the antenatal diagnosis of spina bifida. Lancet 197~ II' 522 28. Harris, 1%., Jennison, R. F., Barson, A. J., Laurence, K. M., Ruoslahti, E., Sepp~l~, M. : Comparison of amniotic-fluid and maternal serum alpha-fetoprotein levels in the early antenatal diagnosis of spina bifida and anencephaly. Lancet 1974 I, 429 29. l"~evin, N.C., Nesbitt, S., Thompson, W.: MyeloeeIe and alpha-fetoprotein in amniotic fluid. Lancet 1973 I, 1383 30. Kjessler, B., Johansson, S. G. O., Sherman, B. S., Gustavson, K.-H., Hultquist, G.: Alphafetoprotein in antenatal diagnosis of congenital nephrosis. Lancet 1975 I, 432 31. Sepp~li~, M., Ruoslahti, E.: Alpha-fetoprotein in maternal serum: a new marker for the detection of fetal distress and intrauterine death. Amer. J. Obstet. Gynee. 115, 48 (1973 b) 32. Chandra, R . K . , Madhavankutty, K., Way, 1%. C.: Serum alpha-fetoprotein levels in patients with cystic fibrosis and their parents and siblings. Brit. reed. J. 1975 I, 714 33. Smith, J. A.: Serum alpha-fetoprotein in cystic fibrosis. Brit. reed. J. 1975 II, 392 34. Wallwork, J . L . , MeFarlane, A., Hingley, S., Ward, A. M.: Serum alpha-fetoprotein in cystic fibrosis. Brit. med. J. 1975 II, 392 35. Brock, D. J. H., Manson, J. C., Raeburn, J. A.: Serum alpha-fetoprotein in cystic fibrosis. Brit. reed. J. 1975 II, 544 36. Fitzsimmons, J. S., Smith, N., Hiller, E. J., Wynne, J. : Serum alpha-fetoprotein in cystic fibrosis. Brit. reed. J. 1975 HI, 544 37. Bdlanger, L., HameI, D., Aubin, G., Bdlanger, M., Bigonesse, G., Dorval, J., Sell, S., Stillman, S., Skelly, H., Scott, J., Harwood, J., Olmsted, N. : Serum alpha-fetoprotein in cystic fibrosis. Brit. reed. J. 1975 IV, 759 38. Kn6pfle, G., 1%otthauwe, H . W . , Lehmann, F.-G.: Serum alpha-fetoprotein in cystic fibrosis. Brit. reed. J. 1975 IV, 459 39. Nishi, S., Hirai, H.: Radioimmunoassay ofal-fetoprotein in hepatoma, other liver diseases and pregnancy. Garm. Monogr. Cancer Res. 14, 79 (1973) 40. Ishii, M.: Radioimmunoassay of ~l-fetoprotein. Gann. Monogr. Cancer 1%es. 14, 89 (1973) 41. Lehmarm, F.-G., Lehmann, D., Martini, G. A. : Isolierung und Kristallisation yon al-Foetoprotein aus mensehlichem Plasma. Clin. chim. Acta 83, 197 (1971) 42. Lehmarm, F.-G., Lehmann, D. : Isolierung und Kristallisation yon ~rFoetoprotein aus einem primi~ren Leberzellkarzinom. Z. klin. Chem. 9, 309 (1971) 43. Maneini, G., Vaerman, J . P . , Carbonara, 0., Heremans, J. F.: A single radial immunediffusion for the immunological quantitation of proteins. Coll. Prof. Biol. fluids I I , 370 (1965) -

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44. Sizaret, P., Breslow, N., Anderson, S. G., Adinolfi, M. C., Buffe, D., Goussev, A., Kohn, J., Krebs, B., Lehmann, F.-G., Martel, N., MeIntire, R., ~ishi, S., Purves, L., Rowe, D. S., Ruoslahti, E.: Collaborative study of a preparation of human cord serum for its use as a reference in the assay of c~1-fetoprotein. J. Biol. Standard. 8, 201 (1975) 45. Lehmann, F.-G., Lehmann, D.: Vergleich verschiedener Methoden zum serologischen Nachweis yon al-Foetoprotein im Serum. Z. klin. Chem. l l , 339 (1973) 46. Sachs, L.: Angewandte Statistik. Berlin-Heidelberg-New York: Springer 1974 47. Craig, J. M., Haddad, A., Shwachman, H.: The pathological changes in the liver in cystic fibrosis of the pancreas. Amer. J. Dis. Child. 98, 357 (1957) 48. Feigelson, J. : Mucoviscidose. P~diatrie 8, 417 (1972) 49. DiSant'Agnese, P. A., Blanc, W. A.: A distinctive type of biliary chirrhosis of the liver associated with cystic fibrosis of the pancreas. Pediatrics 18, 387 (1956) 50. Roberts, W. C.: The hepatic cirrhosis of the cystic fibrosis of the pancreas. Amer. J. Med. 82, 324 (1962) 51. Swift, P. N. : Cystic fibrosis of the pancreas and biliary cirrhosis. Proc. roy. Soc. Med. ~6, 923 (1963) 52. Webster, R., Williams, H.: Hepatic cirrhosis associated with fibrocystic disease of the pancreas. Arch. Dis. Childh. 28, 343 (1953) Dr. G. KnSpfie Prof. Dr. Dr. H. W. Rotthauwe Universit~its-Kinderklinik Adenauerallee 119 D-5300 Bonn Federal Republic of Germany

Prof. Dr. F.-G. Lehmann Medizinische Klinik der Universit~t Marburg MannkopffstraBe 1 D-3550 Marburg Federal Republic of Germany

Serum alpha1-fetoprotein in cystic fibrosis.

Europ. J. Pedia$. 122, 241--248 (1976) 9 by Springer-Verlag 1976 Serum Alphal-Fetoprotein in Cystic Fibrosis G. KnSpfle, H. W. Rotthauwe, and F.-G. L...
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