ICANCER RESEARCH 31, 930—936, July 19711
The Occurrence of a Serum Fetal a , Protein in Developing Mice
and Murine Hepatomas and Teratomas'
Brenda Kahan2 and Lawrence Levine3
Graduate Department of Biochemistry, Brandeis University, Waltham, Massachusetts 02154
SUMMARY
Two fetal-specific serum antigens were demonstrated in
fetal mouse protein extracts and in newborn serum when
examined by immunodiffusion with antiserum to fetal extract.
After dilution of the antiserum, microcomplement fixation
assay was specific for one of these antigens. This antigen had
the electrophoretic mobility of an a@-globulin and a molecular
weight of 65,000 to 70,000. Synthesis of a@ fetal antigen
begins on or before the 10th day of gestation; the total
amount rises to a maximum at 20 days of gestation and falls
sharply after birth. a@ fetal antigen is not present in adult
serum in concentrations greater than 0.0025% that of newborn
serum. Synthesis of a@ antigen is resumed in adult mice
bearing transplantable hepatoma and teratoma tumors. A
clonal in vitro line of testicular teratocarcinoma was found to
synthesize a1 fetal antigen. ct@fetal antigens synthesized by
perinatal mice, hepatomas, and undifferentiated teratoma cells
are serologically identical.
INTRODUcTION
The changing pattern of mammalian serum proteins during
development is presumably a reflection of the sequential gene
activation and repression underlying developmental processes.
New antigens identical with those of adult serum appear
gradually during fetal rat development, i.e. , most of the 14 to
16 adult antigens are present by the time of birth (13).
Concomitantly,
other species unique to the fetal period
disappear after birth.
Specific fetal constituents have been recognized since the
work of Pedersen (21), who described a glycoprotein, fetuin,
comprising nearly 50% of the total protein in fetal calf serum
(4). This level decreased rapidly after birth. Since then, an
a-globulin which is the main component of fetal serum has
been demonstrated in 11 mammalian species (6).
Virtually nothing is known concerning the mechanisms
I Publication
No.
777
from
the
Graduate
Department
of
Biochemistry, Brandeis University. Supported by Research Grant E-222
from the American Cancer Society and Research Grant AI-01940 from
the NIH.
2 Postdoctoral
trainee
supported
by
Training
Grant
NB05241
from
the National Institute of Neurological Diseases and Stroke, NIH.
Present address: The University of Wisconsin, Department of Medical
Genetics, Madison,Wis.53706.
3 American
Cancer
Society
Professor
of
Biochemistry
PRP-21).
Received October 12, 1970; accepted March 1, 1971.
930
(Grant
regulating the appearance and disappearance ofa fetal globulin
during the gestational and postnatal period. In recent years, an
intriguing aspect of a-globulin regulation has been reported. In
cancer cells of the adult organ, genes normally active only
during the fetal period are “derepressed.―
Abelev et aL (2) first
demonstrated that the a-globulin produced by the embryonic
liver Land by the rat yolk sac (7) and possibly the human
placenta (34)] reappears in the serum of adult mice with
hepatomas. This reappearance of fetal protein in the sera of
adults with hepatomas has been subsequently shown to occur
also in humans (3, iS, 32, 33), rats (2, 28), and monkeys (10).
In mice, the fetal protein is synthesized by the cancerous liver
cells (2). Recently, a@ fetal protein has also been found in the
serum of some humans
with teratocarcinomas
(1 , 1 1 , 18, 26).
In this study, we followed the appearance of a fetal protein
in various systems of the mouse. The antigenic identity of the
fetal component synthesized by the developing mouse with
that found in sera of adult mice carrying hepatomas and
certain types of teratomas
is demonstrated.
Systems
synthesizing a-globulin in vitro are of great value for
experimental studies of the nature of the phenomenon in
question. To this end, a clonal in vitro culture of a mouse
testicular teratocarcinoma was investigated and demonstrated
to produce a@ fetal protein.
MATERIALS AND METHODS
Animals and Tumors.
The fetal, postnatal,
and young adult
stages were investigated in Swiss albino nomnbred mice
(Cambridge Research Institute, Cambridge, Mass.). The age of
the fetuses was taken as the number of days after a single
24-hr mating period.
The transplantable hepatoma line BW7756 was obtained
from the Jackson Laboratory, Bar Harbor, Maine. The primary
tumor arose spontaneously in the liver of a strain CS7L/J
mouse and has been transplanted s.c. in 5- to 6-week-old C57/J
male mice. The primary and transplantable teratomas were the
generous gift of Dr. L. C. Stevens. The primary teratomas
arose in the testes of strain TER mice. This inbred strain was
derived from strain 129/Sv and is genetically very similar to it.
Approximately 30% of the males of this strain spontaneously
develop testicular
teratomas
(L. C. Stevens, personal
communication). Adult male mice bearing either unilateral (3
animals) or bilateral (2 animals) tumors weighing 0.6 to 2.0 g
were used. The transplantable teratoma Sv 6965 was derived
from a spontaneous teratoma in the testis of an F2 hybrid
between strains A/He and l29/Sv and has been transplanted
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a1 -Fetal Protein in Developing Mice
@
s.c. in male F1 hybrids between strains A/He and l29/Sv. The
transplantable teratoma OTT 6358 was derived from a strain
C3H/DiSn 6-day embryo grafted to the testis of an adult and
has been transplanted s.c. in adult male mice. The latter tumor
is indistinguishable from transplantable teratomas derived
from spontaneous testicular teratomas (30). The parietal yolk
sac carcinomas, the generous gift of Dr. G. B. Pierce, were
transplanted s.c. into adult female strain 129/J mice. Tumors
removed from the animals averaged 23 g.
Cell Line. A clonal teratocarcinoma line, T1 8-10, has been
described previously (12). Cells were grown in tissue culture in
Falcon Plastic Petri dishes (Falcon Plastics, Oxnard and Los
Angeles, Calif.) or flasks containing Dulbecco's modification
of Eagle's medium supplemented with 15% fetal calf serum. At
each transfer, cells were dissociated for 10 mm in a 4 mM
EDTA (Ca@, MC free) phosphate-buffered
saline solution
containing 2% chicken serum. For the formation of tumors, 3
x 106cells/0.3
mlofculturemedium
wereinoculated
i.p.into
adult 129/J female mice. Many solid tumors formed
subsequently on the peritoneal surface, and small amounts of
ascites fluid were collected.
Collection and Preparation of Samples. Suckling mice were
decapitated to obtain serum. Adult serum was obtained from
mice anesthetized with ether before decapitation. Embryos
were removed with fetal membranes and placenta intact.
Fetuses, including amniotic fluid, and newborns were
homogenized in a glass tissue grinder or electric blender and
treated for 2 to 3 mm in a 250-watt, lO-kc sonic oscillator
(Raytheon Manufacturing Company, Waltham, Mass.) or a
Branson Model W185C sonifier. (Branson Instruments Co.,
Stamford, Conn.). The resulting suspension was centrifuged in
a Sorvall 5534 rotor(Ivan Sorvall, Norwalk, Conn.) at 30,000 X
g for 1 hr at 4°. The supernatant solution was removed and
stored at —20°.
I00
80
C
0
a
‘C
.U
60
C
6)
E
6)
a.
40
E
0
U
20
0.00I
0.0I
Units
@
@
0.1
a1 Fetal
Globulin
Chart 1. Superimposed complement fixation curves obtained with
fetal extract, newborn serum, postnatal serum, and fetal extract
antiserum. 0, 12-day fetal extract; ., 16-day fetal extract;
newborn
serum; £,10-day postnatal serum.
260
240
220
—
20Q
E
)?
ISA
Mouse at
Fetat Gtobut,n
t80
60
se
40
20
—
I0@
i0'
Molecular Weight
Chart 2. Plot of elution volume, Ve,against log molecular weight for
proteins on a Sephadex G-200 column (2 x 100 cm) at pH 7.4.
Tissue culture fluid and some sera were fractionated and
concentrated by neutral (NH4)2 504 ,@recipitation. The liquid
was brought to 40% saturation at 0 by the addition of an
appropriate volume of 100% saturated (NH2 )2 504 solution
adjusted to pH 7. The suspension was allowed to stand for 0.5
hr and was then centrifuged at 30,000 X g for 20 mm. To the
resulting supernatant solution, 1 volume of 100% saturated
neutral (NH@)2SO4 solution was added, bringing the
saturation to 70%. The suspension was allowed to stand for
0.5 hr and was then centrifuged at 30,000 X g for 20 mm. The
supernatant solution was discarded; the pellet was redissolved
in 1 ml of buffer that was used in the complement fixation
assay and dialyzed for 48 hr against 3 changes of 2 liters 0.15
M NaCl, 0.0005
M MgSO4 , and 0.00015
M CaCl2.
Antisera. The soluble protein from strain C3H/HeJ fetuses
freed of extraembryonic components of 11 and 13 days of
gestation was the immunogen. Gestation Day 1 was taken as
the day of vaginal plug appearance. After removal of the
membranes, the fetuses were frozen and thawed, then
homogenized, subjected to sonic oscillation, and centrifuged,
as described above. Antisera were prepared by giving rabbits
injections in the toe pads and i.m. of 0.5 ml of this fetal
extract emulsified in 0.5 ml of complete Freund's adjuvant, 1
injection each week for 3 weeks. The rabbits were bled 1 week
later and given booster doses the following week with 0.4 ml
of immunogen in 0.4 ml of adjuvant i.m. Antiserum Ra4S2C-2
from an animal bled 1 week after receiving booster doses was
used throughout this study.
Immunochemical Methods. Double diffusion plates were
prepared
according
to
Ouchterlony
(20).
Immunoelectrophoresis
in a 1.5% agar gel 0.1 M barbital HC1
buffer (pH 8.6), was performed according to Scheidegger's
method (25). The quantitative complement fixation assay was
performed as described by Levine (17). The diluting buffer
was 0.15 M NaCI-0.OOl M Tris, pH 7.4, containing 1.5 X l0@
M CaCl2 , S X 10
M MgSO4 , and 0.1 BSA.4 Antigens
and
antibodies were heat inactivated at 60°for 20 mm.
°The abbreviation
used is: BSA, bovine serum albumin.
JULY 1971
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931
Brenda Kahan and Lawrence Levine
Estimation of Molecular Weight by Gel Filtration. Molecular
weights were estimated by gel filtration on a Sephadex G-200
column (1 10 x 2 cm) eluted with 0.1 5 M NaC1-0.0l M Tris
buffer, (pH 7.4), at a flow rate of 20 mi/hr. The column was
calibrated with cytochrome c (Mann Research, New York, N.
Y.), BSA (Armour and Company, Chicago, Ill.), and fumarase
(Calbiochem., Los Angeles, Calif.) standards. Protein standards
were located by absorbance at 413 mp (cytochrome c) or 280
m,.t (BSA and fumarase). The position of the fetal antigen peak
was determined after quantitation by complement fixation.
2000
1600
RESULTS
E
a,
It was first necessary to demonstrate that the fetal
extract antiserum was capable of detecting fetal-specific
antigens and that one of these antigens had the characteristics
of a-globulin previously described as being present in the fetal
sera of mice and other species. This was accomplished in the
following experiments.
Antibodies directed toward adult serum components were
effectively eliminated from the agar diffusion experiments by
incorporation of 10% adult serum in the agar. Under these
conditions, no band of precipitation was observed with the
antiserum and normal adult serum. However, the fetal extract
200
U)
E
a,
U)
C
800
400
I00
0
4
10
8
12
16
20
24
28
Days After Birth
Chart 4. The disappearance of a@fetal globulin in the serum of mice
during the postnatal period. Serum concentrations for days after birth
are values derived from pooled sera of 2 to 22 mice: o, each point
represents pooled scm of all mice in 2 litters; ., each point represents
10 to 11 mice from Set 1 of 6 pooled litters; @,
each point represents
pooled sera of 2 to 9 mice from Set 2 of 6 pooled litters; A, each point
1000
900
800
represents pooled sera of 14 mice from Set 3 of 6 pooled litters.
700
a
E
used as immunogen or aqueous extracts of 12-, 14-, 16-, and
18-day-old fetuses and newborn serum all gave 2 bands of
precipitation (1 major and I minor band) with this antiserum.
One of these antigens (the minor one) disappears from the
serum by the 3rd day after birth. In serum obtained from
18-day-old mice, the major band is stifi observed but is much
weaker. At 28 days, the antigen has disappeared, as measured
by gel diffusion.
In the complement fixation assay, the antiserum at a
dilution of 1/500 was monospecific even without absorption
with adult serum. Complement fixation curves obtained with
12- and 16-day-old fetal extracts and with newborn and
postnatal serum (10 days) reached the same maximum fixation
and could be superimposed in both antibody and antigen
excess regions (Chart 1). There was no complement fixation
600
C
4
500
a,
U)
C
D
400
300
200
100
10
12
4
16
18
20
Days of Gestation
I
3
5
Days After Birth
Chart 3. Soluble protein concentrations of a@fetal globulin in the
mouse as a function of age. The value expressed for a given day
represents the average amount per animal from 2 litters; each litter of 5
to 14 mice was pooled and assayed separately.
932
with
adult
serum.
The
anticomplementary
properties
of adult
serum prevented the assay of specific antigen-antibody
reactions at high concentrations. A 40 to 70% (NH@)2504 cut
of newborn and adult serum mixtures precipitated over 95% of
the fetal antigen and removed much of the complement
inhibiting
activity.
This
40
to
70% (NH@)2SO@
CANCER RESEARCH
VOL. 31
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a1 -Fetal Protein in Developing Mice
cut of adult serum reconstituted in buffer was negative by
complement fixation assay at a dilution of 1/2.5 . The fetal
antigen is therefore
not present in adult serum in
concentrations greater than 0.0024% that of newborn serum.
Early postnatal serum contains the fetal specific antigen in
high concentration. On immunoelectrophoresis,
this antigen
migrates slightly behind albumin to a position identifying it as
an a1 -globulin protein.
The molecular weight of a1 fetal antigen was estimated by
G-200 gel filtration of hepatoma serum. As shown below, the
a1 -globulin in newborn serum and that present in the serum of
hepatoma-bearing mice are serologically identical. The elution
profile of a1 -globulin, which was estimated by complement
fixation, was indistinguishable from that of BSA (Chart 2). A
molecular weight estimate for mouse a1 fetal protein of
65,000 to 70,000 agrees well with that previously determined
by gel filtration for human (5, 9) and rat (I 6) a1 fetal protein
as well as with a molecular weight of 64,800 calculated by the
Svedberg method for rat fetal postalbumin (14). These
proteins thus appear to be somewhat different from fetuin,
which has a molecular weight of 48,400 (27).
a1 Antigen in Developing Fetuses and Postnatal Animals.
The appearance of a1 antigen in the soluble protein fraction of
fetuses after 10 to 20 days of gestation and its disappearance
in postnatal serum and soluble protein was followed by
complement fixation assay.5
The data indicate that a1 antigen was present in the earliest
fetuses that were obtained. The amount of antigen present
increased by over 4 logs in 4 days and continued to rise
sharply until 20 days of gestation (Chart 3). After birth, there
was an immediate and sharp decrease in the soluble protein
fraction and serum antigen concentrations, which fell to
undetectable levels by the 28th day after birth (Chart 4).
a1 Antigen in Maternal Sera. a1 antigen was detected by
complement fixation in the sera of adult females during
gestation. The concentrations in the sera of 13- to 18-day
pregnant mice were approximately 2% of the total amount per
embryo of the same gestation age. Fetal antigen could not be
detected in some of these same sera before (Nit, )2 504
fractionation due to the anticomplementary properties of the
serum at concentrations higher than 1/150.
a1 Antigen in Sera of Adult Mice with Hepatomas and
Teratomas. The a1 antigen synthesized in adult hepatoma and
teratoma tumor-bearing animals is immunologically identical
with that in perinatal mice (Fig. 1). The identity of the antigen
in the 3 cases is further confirmed by the fact that a1 antigen
from each source gave identical complement fixation curves,
each of which could be eliminated by throwing the system
S A 0.01
unit
of
a@ antigen
is defined
as the
amount
of antigen
into antigen excess with the calculated amount of serum from
1 of the other sources.
The data in Chart 5 show the increase of a1 antigen in the
serum of adult mice as a function of time after transplantation
of hepatoma BW7756. Mice receiving inoculations on Day 0
had concentrations
of circulating a1 antigen on Day 28
exceeding by over a factor of 2 those of newborn serum.
Mice with transplantable teratomas had much lower levels
of a1 antigen than mice with hepatomas (Table 1). The
complement fixation curves of a1 fetal antigen found in the
ascites
fluid
and/or
serum
of each mouse
examined
with a
transplantable teratoma could again be superimposed on a
single curve, demonstrating identity of the antigen. Cells of the
clonal teratoma line, T1 @,8-10, synthesized a1 protein in vitro
(Chart 6) as well as in vivo (Table 1).
In contrast, a1 fetal protein was not present in mice with
primary teratomas. The minimum concentration of a1 fetal
antigen that could have been detected in sera of 5 mice with
primary testicular teratomas or in fluid exudates from 3 of
these tumors ranged between 0.12 and 0.4 unit/ml of
undiluted sample and was determined by the variable
anticomplementary
properties of the fractionated sera and
fluid exudates. However, in no case could any of these fluids
have had a concentration of more than 0.4 unit/ml, a value
below the range (0.7 to 18.6 units/mI) of a1 fetal protein
present in transplantable teratoma fluids. Primary teratoma
0.000
000
E
a,
U)
E
l00
a
U)
C
D
I0
that
gives half-maximum fixation in the region of antibody excess. Thus, if
half-maximum fixation in the region of antibody excess is obtained
with a 1/10,000 dilution of a serum or a fetal protein extract, this
serum or extract contains 100 units/ml. In this report, 1 unit of a,
antigen is assumed to be equal to 1 @gof a1 antigen, an assumption
that has been shown to be valid in an analogous human fetal a,
0
immune system and closely approximated in several other protein
immune systems studied by C' fixation (L. Levine, unpublished data).
Although the 2 values may prove not to be exact, they provide
convenient references for evaluation of our data.
4
8
2
Days After
6
20
24
28
32
Transplantation
Chart 5. Increasingserum concentrations of a, fetal globulin in mice
receiving hepatoma
BW7756 as a function
of days after transplantation.
JULY 1971
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933
Brenda Kahan and Lawrence Levine
‘@
c:!I
1@1
Ii,
r@1
Fig. 1. Double diffusion in agar containing 10% adult serum. A, fetal extract antiserum (undiluted); B, 18-day postnatal serum (undiluted); C,
hepatoma serum (diluted 1/100); D, teratoma 6358 serum (undiluted).
100
Table 1
@
Serum and ascites concentration of , antigen in mice
with transplantable teratomas
AnimalTeratomaNo.SourceUnits/ml0TT6358ISerum13.2T,7
C
0
80
0
‘C
8—101Ascites11.2T,
8—102Ascites18.6T,78—lO2Serum6.30TT69651Serum0.7
.@
60
C
a)
E
a)
a.
E
40
0
U
fluids therefore do not contain a1 fetoglobulin or do so at
lower concentrations than do transplantable teratoma fluids.
20
DISCUSSION
@
@
Primary and transplantable teratomas of mice have been
extensively examined by Stevens (29), and Stevens and
Hummel (3 1). The primary teratomas of older mice are usually
benign and characteristically consist of nonproliferating, fully
differentiated tissues, whereas the transplantable teratomas
invariably contain dividing, undifferentiated
stem cells.
Transplantable
tumors
may
also contain
immature,
differentiating tissues that are dividing. a1 fetal protein may,
therefore, be synthesized by all dividing teratoma cells or only
by the undifferentiated
stem cell type. These possibilities
could be differentiated if a form of teratoma were available
that consisted of only differentiated cells and also was capable
of rapid division. The parietal yolk sac carcinoma of Pierce and
Dixon (22) was originally derived from a transplantable
testicular teratoma of strain 129/J mice. The cells of this
934
0.001
0.01
Units
0.1
a1 Fetal
Globulin
Chart 6. Superimposed complement fixation curves of concentrated
tissue culture medium of T, 8-10 cells in vitro: o, T, 8-10 tissue
culture
serum.
medium;
@,
T, ., 8-10 tissue culture
medium;
., 3-day postnatal
tumor are not multipotential
like the stem cells of
differentiating teratomas but produce only hyaline parietal
yolk sac substance. The sera of 2 mice bearing parietal yolk sac
carcinoma were negative for
fetal globulin ; the lower limit
of detection in the complement fixation assay was 0.1 unit/ml.
This result is compatible with the hypothesis that a1 fetal
globulin is synthesized only by the undifferentiated
or
embryonal carcinoma cells of the teratoma. Thus, no primary,
CANCER RESEARCH VOL. 31
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a1 -FetaiProtein
@
@
well-differentiated
teratoma
produced a1 fetal protein,
whereas 100% of transplantable teratomas examined did.
However, the most cogent support of this hypothesis is the
fact that T1 8-10 cells synthesize a1 fetal antigen in vitro and
in vivo. These cells grow in culture as a compact monolayer of
undifferentiated epithelial cells. In addition, T1 8-10 is a
well-characterized clonal line of teratoma cells producing at
this time tumors composed almost entirely of undifferentiated
embryonal carcinoma cells; a very few areas of parietal yolk
sac differentiation were found in sectioned tumors. [The
clonal cells of earlier generations were multipotential;
well-differentiated teratoma, which did not include liver tissue,
formed after injection of these clonal cells into mice (12)].
The synthesis of a1 protein by undifferentiated teratoma
cells rules out explanations previously offered for the finding
of a1 in human teratomas of either synthesis by tumor cells
determined to become embryonic liver cells or of an effect of
the tumor on the host liver causing a resumption of a1
synthesis (1).
It appears that synthesis of a1 fetal globulin begins on or
before the 10th day of gestation. The total amount of a1
protein reaches a maximum in 20-day-old fetuses and falls
rapidly after birth. The rapid inhibition of the synthesis of a1
fetal globulin at birth parallels similar findings for this antigen
in man (where inhibition slightly precedes birth) (5) and in
rats (7) and that previously determined qualitatively by
Ouchterlony
assay in mice (2). The mechanism of this
regulation is unknown.
The reactivation of a fetal liver gene in mouse hepatoma
cells closely parallels the fmdings by Gold and Freedman (8)
of an antigen present in organs of the fetal digestive tract in
cancer of the corresponding adult organs. Some Morris
hepatomas have been found to synthesize an aldolase isozyme
characteristic of fetal liver (19, 24). These studies suggest that
an entire spectrum of fetal-characteristic gene products should
be investigated further to evaluate the hypothesis that
carcinogenic transformation
results in the elaboration of
certain states resembling those of the fetal organism (23).
The normal counterparts of murine embryonal teratoma
cells are either the primordial germ cells or very early
embryonic cells. It is not known whether these (normal) cells
are capable of producing a1 fetal antigen. On the other hand,
synthesis of a1 fetal protein by teratoma cells may be the
production of an “ectopic―
substance by cancerous cells.
However, teratoma cells are unique among cancer cells in their
multipotentiality. Synthesis of a1 fetal protein may, therefore,
be another manifestation of their peculiar embryonic-like
nature.
in Developing Mice
2. Abelev, G. I., Perova, S. D., Khramkova, N. I., Postnikova, Z. A.,
and hlin,
I. S. Production of Embryonal co-Globulin by
Transplantable Mouse Hepatomas. Transplantation, 1: 174—180,
1963.
3. Alpert, M. E., Uriel, J., and de Nechaud, B. Alpha, Fetoglobulin in
the Diagnosis of Human Hepatoma. New Engl. J. Med., 278:
984—986, 1968.
4. Bergmann, F. H., Levine, L., and Spiro, R. G. Fetuin.
Immunochemistry and Quantitative Estimation in Serum. Biochim,
Biophys. Acta, 58: 41—51, 1962.
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7-Globulin in the Human Conceptus. J. Clin. Invest., 45:
1826—1838,1966.
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Several Mammals and Their Relation to Human ot-Fetoprotein.
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1010—1016,1967.
8. Gold, P., and Freedman, S. 0. Specific Carcinoembryonic Antigens
of the Human Digestive Tract. J. Exptl. Med., 122: 467—481,
1965.
9. Houlték,
J., Masopust, G., Kithier, K., and RádI,J. Hepatocellular
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Fetoprotein. J. Pediat., 72: 186—193,1968.
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M. G. a-Fetoprot@in in Monkeys with Hepatoma. J. NatI. Cancer
Inst., 42: 1035—1044, 1969.
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ot-Fetoprotein (otFP) in Cancer Patients. Clin. Res., 1 7: 403, 1969.
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Clonal in Vitro Cultures of Mouse Testicular Teratoma. J. Natl.
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Rat Studied by Immunoelectrophoresis.
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Biochim. Biophys. Acta,
14. Kirsh, I., Wise, R., and Oliver, I. I. Post-Albumin,a Foetal-specific
Rat Plasma Protein. Purification, Physicochemical and
Immunological Studies. Biochem. J., 102: 763—766,1967.
15. Kithier, K., Houstek, G., Masopust, J., and Radl, G. Occurrence of
a Specific Foetal Protein in a Primary Liver Carcinoma. Nature,
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16. Kithier, K., and Prokes, J. Fetal a, -Globulin of Rat Serum.
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Handbook of Experimental Immunology, pp. 707—719.Oxford:
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ACKNOWLEDGMENTS
We thank Dr. James Zuckerman for helpful discussions of and
interest in this work.
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CANCER RESEARCH
VOL. 31
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The Occurrence of a Serum Fetal α1 Protein in Developing Mice
and Murine Hepatomas and Teratomas
Brenda Kahan and Lawrence Levine
Cancer Res 1971;31:930-936.
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