- Email: [email protected]
The effect of the intrauterine contraceptive device on the prevalence of morphologic abnormalities in human spontaneous abortions
CONTRACEPTION
THE EFFECT OF THE INTRAUTERINE CONTRACEPTIVE DEVICE ON THE PREVALENCE OF MORPHOLOGIC ABNORMALITIES IN HUMAN SPONTANEOUS ABORTIONS
L. H. HONOR;* OF LABORATORIES, GRACE GENERAL HOSPITAL and OF PATHOLOGY, MEMORIAL UNIVERSITY OF NEWFOUNDLAND ST. JOHN'S, NEWFOUNDLAND, CANADA
DEPARTMENT DIVISION
*NEW ADDRESS:
Dr. Louis H. Honor6 Provincial Laboratory Pathology Department University of Alberta Edmonton, Alberta Canada T6G 252
of Public Health
ABSTRACT
Spontaneously aborted conceptuses from IUD-wearing patients and prior IUD users were examined pathologically to determine the prevalence of morphologic abnormalities, which had previously been shown to correlate with cytogenetically proven heteroploidy in the conceptus. These abnormalities were never seen in the IUD-associated abortuses while occurring with the same frequency in abortuses obtained from prior IUD users and from non-users. It is concluded that the IUD by itself is responsible for the absence of heteroploidy-correlated structural abnormalities in spontaneous abortuses conceived with the IUD -in situ. Relevant mechanisms underlying this IUD effect are discussed.
Accepted
for publication
December
JANUARY 1980VOL. 21 NO. 1
26, 1979
47
CONTRACEPTION
INTRODUCTION A combined pathologic and cytogenetic examination of spontaneously aborted conceptuses led to the recognition of certain morphologic abnormalities, which correlated highly with cytogenetically proven heteroploidy in the conceptus (1). When this method was used to study 514 spontaneously aborted and therapeutically evacuated conceptuses from healthy women wearing the IUD, these heteroploidy-correlated structural abnormalities were consistently absent (2). In the same study population these abnormalities were detected pathologically in 0.83% of 5779 therapeutic abortions unrelated to concurrent IUD use and the overall prevalence of cytogenetically documented heteroploidy in IUD-unrelated spontaneous abortions was 46.7% (3). It was concluded that the IUD suppressed the development of these abnormal conceptuses and by inference it was suggested that the IUD significantly affected the occurrence of heteroploidy in human pregnancies (2). The responsibility of the IUD for these effects was further tested by comparing the prevalence of heteroploidy-correlated structural abnormalities in spontaneous abortuses obtained from concurrent and prior IUD users.
MATERIALS
AND METHODS
Tha author personally examined tissues from 617 spontaneous abortions, using a standardized procedure involving the following steps: qualitative and quantitative assessment of the chorionic sac (if present) and its contents; external evaluation of the embryo/fetus for growth disorganization (4); internal scrutiny of the conceptus for anomalies; and systematic examination of the decidua and placenta (washed clean and floated in saline) for hemorrhage and villous agglutination, clubbing and macrocystic change. These tissues were processed -in toto or extensively sampled for histology. Pathologic interpretation, only attempted when adequate tissue was available for study, was based on the detection of heteroploidy-correlated gross and microscopic abnormalities in the placenta and in the embryo/ fetus, if available. In the absence of these abnormalities a diagnosis When these abnormalities were of "suggestive of diploidy" was made. found, they were interpreted in terms of the chromosomal anomaly associated with the abortions and an inferential diagnosis of "suggestive of heteroNo cytogenetic studies were done ploidy, i.e. triploidy etc." was made. on these abortuses. The patients' charts were reviewed after the pathologic diagnosis was made, and special attention was paid to associated medical, surgical or reproductive disorders, contraceptive practices and relevant details Use of the IUD in the past or at the about the abortion under study. time of the abortion was recorded.
48
JANUARY
1980 VOL. 21 NO. 1
CONTRACEPTION
RESULTS Of these 617 spontaneous aborters, 22 had an IUD -in situ at the time of conception and abortion and had been using the device (Lippes Loop: 20 and Copper-T: 2) for 6 months to 7 years uninterruptedly, with the majority having worn it for 2-3 years. The IUD (Lippes Loop: 19, Dalkon Shield: 3 and Copper-T: 1) had been worn in the past by 23 of these spontaneous aborters and had been removed 2 weeks to 6 years before pregnancy occurred and ended in the spontaneous abortion studied. In a 24-year-old primigravida, the IUD had been removed during the cycle when pregnancy occurred, i.e. on day 12 of a fairly regular 28-to 30-day cycle. Complete pathologic evaluation, including a comment on the "karyotype" of the abortus, was possible in 422 of the 617 cases examined (68.4%). Of the 22 concurrent IUD users,17 passed enough tissue for interpretation; none of these cases showed heteroploidy-correlated morphologic abnormalities and were diagnosed as "suggestive of diploidy". The menstrual ages of these abortions ranged from 56 to 155 days, with 21.9% (7/22) being less than 12 weeks. Of the 23 prior IUD users,17 passed enough tissue for interpretation: 7 of these abortuses were structurally normal and diagnosed as "suggestive of diploidy"; 12 showed heteroploidy-correlated morphologic abnormalities, 9 being labelled as "suggestive of trisomy", 2 "suggestive of triploidy" and 1 "suggestive of monosomy X". The menstrual ages of these abortions ranged from 63 to 156 days, with 69.6% (16/23) being less than 12 weeks. The abortuses, obtained from the prior IUD users and from the nonusers, showed no statistical difference (p >O.lOO) in the overall prevalence and distribution of the various types of heteroploidycorrelated morphologic abnormalities, In contrast the concurrent IUD users and the prior IUD users were statistically different (p(O.010) as regards the prevalence of these structural abnormalities in their abortuses.
DISCUSSION This study is based on uncoventional methodology which demands critical evaluation. The pathological technique used is an extension of the original work of Philippe and Boug (5) and was developed specifically to detect morphologic abnormalities in abortuses that correlated with the broad categories of reproductive heteroploidy (1). Once the phenotypic correlates of these karyotypic anomalies were systematized and objectivated, it was possible to deduce the likely "karyotype" of an abortus from its morphology with a high level of accuracy, i.e., an overall 94% accuracy in differentiating cytogenetically proven diploid and heteroploid abortuses and an 80% accuracy in properly categorizing the type of heteroploidy present (2). The method, which attempts to bridge the gap between phenotype and karyotype, needs further validation
JANUARY
1980 VOL. 21 NO. 1
49
CONTRACEPTION
and refinement and so far preliminary support its basic validity (6).
results from a collaborative
study
In confirmation of previous findings (2), the abortuses, obtained from women currently wearing the IUD, are consistently free of detectable hetercploid-correlated morphologic abnormalities, which occur with the It is concluded that same frequency in prior IUD users and non-users. the IUD by itself is responsible for the selective disappearance of abortuses with such abnormalities. Pending cytogenetic validation or refutation, it is suggested by inference that the IUD preferentially This finding suppresses the development of heteroploid conceptuses. is consistent with the following observations: 1) severe growth disorganization of embryo/fetus is distinctly less common in IUD-associated abortions (3); 2) chromosomal anomalies have not been reported in offspring born with the IUD -in situ (7), though this problem has not been systematically investigated; and 3) no significant teratogenicity of the IUD has been demonstrated (8,9). This study sheds no light on the factors underlying this IUD effect. It is suggested that the IUD, which alters the uterine milieu (lO,ll), prevents the development of conceptuses with heteroploid-correlated abnormalities by at least three mechanisms: 1) by preferentially destroying these basically less viable zygotes (12); 2) by selectively eliminating the abnormal spermatozoa responsible for heteroploidy (13-X); and 3) by interfering with sperm migration (16) and reducing the likelihood of polyploidy secondary to excessive tubal flooding with spermatozoa (17). With the available technology these suggestions are directly amenable to experimental and clinical testing. This study cannot answer the question of whether IUD-associated conceptuses, subclinically aborted, also fail to exhibit detectable heteroploidy-correlated abnormalities. Acquisition of such data would help to define when in early gestation the IUD exerts its deleterious effect on these abnormal embryos. Experimental techniques are available (18) for a systematic attack on the problem. Clinically, with the advent of sensitive pregnancy tests (19-21), it is now possible early to identify women pregnant with the IUD -in situ. Should any of these women desire early therapeutic abortion, their abortuses could be karyoThere is clearly a need for concerted clinical, cytogenetic and typed. morphologic investigations in this field of contraception.
50
JANUARY
1980 VOL. 21 NO. 1
CONTRACEPTION
REFERENCES
1.
Honorg, L.H., Dill, F.J. and Poland, B.J. Placental morphology Teratology 14: abortuses with normal and abnormal karyotypes. 151-166 (1976).
2.
Honor;, L.H. Early human pregnancy with the intrauterine contraceptive device -___ in situ: Incidence of heteroploidy, Teratology 20: 3-6 (1979).
3.
Poland,
4.
Ploand, B.J. and Miller, J.R. Effect of karyotype on zygotic development. In: Comptes Rendus du Colloque: Les Accidents Chromosomiques de la Reproduction (A. Bou; and C. Thibault,Editors) INSERM, Paris, 1973, p 111-118.
5.
Philippe, E. and Boue, J.G. Placenta et aberrations au tours des avortements spontane/s. Presse Med 78:
6.
Warburton, Teratology
7.
Rosenfield, assessment.
B.J.
Personal
Communication
D. Heteroploidy 20: 2 (1979). A.
(1978).
in IUD-associated
Oral and intrauterine
Am J Obstet Gynecol 132:
in
chromosomiques 641-646 (1970).
pregnancy
(Editorial).
contraception: a 1978 risk 92-106 (1978).
8.
Tietze, C. Evaluation of intrauterine devices. Ninth Progress Report, Cooperative Statistical Program, Studies in Family Planning No. 55 (1970).
9.
Chang, C.C. and Tatum, J.H. Absence of teratogenicity of intrauterine copper wire in rats, hamsters and rabbits. Contraception 7: 413-434 (1973).
10.
Tamaya, T., Nakata, Y., Ohno, Y., Nioka, S., Furuta, M. and Okada, H. The mechanism of action of the copper IUD. Fertil Steril 27: 767-772 (1976),
11.
Parr, E.L. and Shirley, R.L. Embryotoxicity of leukocyte extracts and its relationship to intrauterine contraception. Fertil Steril 27: 1067-1077 (1976).
12.
Cur:, S., Boug, J.G. and Bou&, A. Growth characteristics of human embryonic cell lines with chromosomal anomalies. Biomedicine 21: 233-236 (1974).
13.
Jonasson, J., Therkelsen, A.J., Lauritsen, J.G. and Lindsten, J. Origin of triploidy in human abortuses. Hereditas 71: 168-171 (1972).
14.
Bomsel-Helmreich, 0. Aging of gametes, heteroploidy death. Int J Gynecol Obstet 14: 98-104 (1976).
JANUARY
1980 VOL. 21 NO. 1
and embryonic
51
CONTRACEPTION
15.
Lauritsen, in Trisomy
16.
Tredway, C.R., Umezaki, C.U., Mishell, D.R., Jr. and Settlage, Effect of intrauterine devices on sperm transport in the human being. Am J Obstet Gynecol 123: 734-735 (1975).
17.
Hunter, R.H.F. and L'Englise, P.C. Polyspermic fertilization following tubal surgery in pigs with special reference to the role of the isthmus. J Reprod Fertil 24: 233-246 (1971).
18.
In vitro Hurst, P.R., Jefferies, K., Dawson, K.J. and Eckstein, P. -development of preimplantation embryos recovered from IUD-bearing J Reprod Fertil 54: 413-417 (1978). mice.
19.
Roy, S., Klein, T.A., Scott, J.Z., Rletsky, Diagnosis of pregnancy with a radioreceptor Gynecol 50: 401-406 (1977).
20.
A simple and sensitive nonradioactive Banik, U.K. and Givner, M.L. method for the detection of urinary human chorionic gonadotropin and diagnosis of early human pregnancy. I. Multiple-unit test. Fertil Steril 32: 420-425 (1979).
21.
Banik, U.K., Hirsch, M.A., Irvine, D.S., Krupey, J., Hurwitz, A., Singh, K., Wetzel, .I., and Givner, M.L. A simple and sensitive nonradioactive method for the detection of urinary human chorionic gonadotrophin and diagnosis of early human pregnancy. II. Singleunit test. Fertil Steril 32: 426-432 (1979).
52
J.B. and Friedrich, U. Origin of the extra chromosome 16. Clin Gen 10: 156-160 (1976). C.S.
O.A. and Mishell, D.R., Jr. assay for hCG. Obstet
JANUARY
1980 VOL. 21 NO. 1
THE EFFECT OF THE INTRAUTERINE CONTRACEPTIVE DEVICE ON THE PREVALENCE OF MORPHOLOGIC ABNORMALITIES IN HUMAN SPONTANEOUS ABORTIONS
L. H. HONOR;* OF LABORATORIES, GRACE GENERAL HOSPITAL and OF PATHOLOGY, MEMORIAL UNIVERSITY OF NEWFOUNDLAND ST. JOHN'S, NEWFOUNDLAND, CANADA
DEPARTMENT DIVISION
*NEW ADDRESS:
Dr. Louis H. Honor6 Provincial Laboratory Pathology Department University of Alberta Edmonton, Alberta Canada T6G 252
of Public Health
ABSTRACT
Spontaneously aborted conceptuses from IUD-wearing patients and prior IUD users were examined pathologically to determine the prevalence of morphologic abnormalities, which had previously been shown to correlate with cytogenetically proven heteroploidy in the conceptus. These abnormalities were never seen in the IUD-associated abortuses while occurring with the same frequency in abortuses obtained from prior IUD users and from non-users. It is concluded that the IUD by itself is responsible for the absence of heteroploidy-correlated structural abnormalities in spontaneous abortuses conceived with the IUD -in situ. Relevant mechanisms underlying this IUD effect are discussed.
Accepted
for publication
December
JANUARY 1980VOL. 21 NO. 1
26, 1979
47
CONTRACEPTION
INTRODUCTION A combined pathologic and cytogenetic examination of spontaneously aborted conceptuses led to the recognition of certain morphologic abnormalities, which correlated highly with cytogenetically proven heteroploidy in the conceptus (1). When this method was used to study 514 spontaneously aborted and therapeutically evacuated conceptuses from healthy women wearing the IUD, these heteroploidy-correlated structural abnormalities were consistently absent (2). In the same study population these abnormalities were detected pathologically in 0.83% of 5779 therapeutic abortions unrelated to concurrent IUD use and the overall prevalence of cytogenetically documented heteroploidy in IUD-unrelated spontaneous abortions was 46.7% (3). It was concluded that the IUD suppressed the development of these abnormal conceptuses and by inference it was suggested that the IUD significantly affected the occurrence of heteroploidy in human pregnancies (2). The responsibility of the IUD for these effects was further tested by comparing the prevalence of heteroploidy-correlated structural abnormalities in spontaneous abortuses obtained from concurrent and prior IUD users.
MATERIALS
AND METHODS
Tha author personally examined tissues from 617 spontaneous abortions, using a standardized procedure involving the following steps: qualitative and quantitative assessment of the chorionic sac (if present) and its contents; external evaluation of the embryo/fetus for growth disorganization (4); internal scrutiny of the conceptus for anomalies; and systematic examination of the decidua and placenta (washed clean and floated in saline) for hemorrhage and villous agglutination, clubbing and macrocystic change. These tissues were processed -in toto or extensively sampled for histology. Pathologic interpretation, only attempted when adequate tissue was available for study, was based on the detection of heteroploidy-correlated gross and microscopic abnormalities in the placenta and in the embryo/ fetus, if available. In the absence of these abnormalities a diagnosis When these abnormalities were of "suggestive of diploidy" was made. found, they were interpreted in terms of the chromosomal anomaly associated with the abortions and an inferential diagnosis of "suggestive of heteroNo cytogenetic studies were done ploidy, i.e. triploidy etc." was made. on these abortuses. The patients' charts were reviewed after the pathologic diagnosis was made, and special attention was paid to associated medical, surgical or reproductive disorders, contraceptive practices and relevant details Use of the IUD in the past or at the about the abortion under study. time of the abortion was recorded.
48
JANUARY
1980 VOL. 21 NO. 1
CONTRACEPTION
RESULTS Of these 617 spontaneous aborters, 22 had an IUD -in situ at the time of conception and abortion and had been using the device (Lippes Loop: 20 and Copper-T: 2) for 6 months to 7 years uninterruptedly, with the majority having worn it for 2-3 years. The IUD (Lippes Loop: 19, Dalkon Shield: 3 and Copper-T: 1) had been worn in the past by 23 of these spontaneous aborters and had been removed 2 weeks to 6 years before pregnancy occurred and ended in the spontaneous abortion studied. In a 24-year-old primigravida, the IUD had been removed during the cycle when pregnancy occurred, i.e. on day 12 of a fairly regular 28-to 30-day cycle. Complete pathologic evaluation, including a comment on the "karyotype" of the abortus, was possible in 422 of the 617 cases examined (68.4%). Of the 22 concurrent IUD users,17 passed enough tissue for interpretation; none of these cases showed heteroploidy-correlated morphologic abnormalities and were diagnosed as "suggestive of diploidy". The menstrual ages of these abortions ranged from 56 to 155 days, with 21.9% (7/22) being less than 12 weeks. Of the 23 prior IUD users,17 passed enough tissue for interpretation: 7 of these abortuses were structurally normal and diagnosed as "suggestive of diploidy"; 12 showed heteroploidy-correlated morphologic abnormalities, 9 being labelled as "suggestive of trisomy", 2 "suggestive of triploidy" and 1 "suggestive of monosomy X". The menstrual ages of these abortions ranged from 63 to 156 days, with 69.6% (16/23) being less than 12 weeks. The abortuses, obtained from the prior IUD users and from the nonusers, showed no statistical difference (p >O.lOO) in the overall prevalence and distribution of the various types of heteroploidycorrelated morphologic abnormalities, In contrast the concurrent IUD users and the prior IUD users were statistically different (p(O.010) as regards the prevalence of these structural abnormalities in their abortuses.
DISCUSSION This study is based on uncoventional methodology which demands critical evaluation. The pathological technique used is an extension of the original work of Philippe and Boug (5) and was developed specifically to detect morphologic abnormalities in abortuses that correlated with the broad categories of reproductive heteroploidy (1). Once the phenotypic correlates of these karyotypic anomalies were systematized and objectivated, it was possible to deduce the likely "karyotype" of an abortus from its morphology with a high level of accuracy, i.e., an overall 94% accuracy in differentiating cytogenetically proven diploid and heteroploid abortuses and an 80% accuracy in properly categorizing the type of heteroploidy present (2). The method, which attempts to bridge the gap between phenotype and karyotype, needs further validation
JANUARY
1980 VOL. 21 NO. 1
49
CONTRACEPTION
and refinement and so far preliminary support its basic validity (6).
results from a collaborative
study
In confirmation of previous findings (2), the abortuses, obtained from women currently wearing the IUD, are consistently free of detectable hetercploid-correlated morphologic abnormalities, which occur with the It is concluded that same frequency in prior IUD users and non-users. the IUD by itself is responsible for the selective disappearance of abortuses with such abnormalities. Pending cytogenetic validation or refutation, it is suggested by inference that the IUD preferentially This finding suppresses the development of heteroploid conceptuses. is consistent with the following observations: 1) severe growth disorganization of embryo/fetus is distinctly less common in IUD-associated abortions (3); 2) chromosomal anomalies have not been reported in offspring born with the IUD -in situ (7), though this problem has not been systematically investigated; and 3) no significant teratogenicity of the IUD has been demonstrated (8,9). This study sheds no light on the factors underlying this IUD effect. It is suggested that the IUD, which alters the uterine milieu (lO,ll), prevents the development of conceptuses with heteroploid-correlated abnormalities by at least three mechanisms: 1) by preferentially destroying these basically less viable zygotes (12); 2) by selectively eliminating the abnormal spermatozoa responsible for heteroploidy (13-X); and 3) by interfering with sperm migration (16) and reducing the likelihood of polyploidy secondary to excessive tubal flooding with spermatozoa (17). With the available technology these suggestions are directly amenable to experimental and clinical testing. This study cannot answer the question of whether IUD-associated conceptuses, subclinically aborted, also fail to exhibit detectable heteroploidy-correlated abnormalities. Acquisition of such data would help to define when in early gestation the IUD exerts its deleterious effect on these abnormal embryos. Experimental techniques are available (18) for a systematic attack on the problem. Clinically, with the advent of sensitive pregnancy tests (19-21), it is now possible early to identify women pregnant with the IUD -in situ. Should any of these women desire early therapeutic abortion, their abortuses could be karyoThere is clearly a need for concerted clinical, cytogenetic and typed. morphologic investigations in this field of contraception.
50
JANUARY
1980 VOL. 21 NO. 1
CONTRACEPTION
REFERENCES
1.
Honorg, L.H., Dill, F.J. and Poland, B.J. Placental morphology Teratology 14: abortuses with normal and abnormal karyotypes. 151-166 (1976).
2.
Honor;, L.H. Early human pregnancy with the intrauterine contraceptive device -___ in situ: Incidence of heteroploidy, Teratology 20: 3-6 (1979).
3.
Poland,
4.
Ploand, B.J. and Miller, J.R. Effect of karyotype on zygotic development. In: Comptes Rendus du Colloque: Les Accidents Chromosomiques de la Reproduction (A. Bou; and C. Thibault,Editors) INSERM, Paris, 1973, p 111-118.
5.
Philippe, E. and Boue, J.G. Placenta et aberrations au tours des avortements spontane/s. Presse Med 78:
6.
Warburton, Teratology
7.
Rosenfield, assessment.
B.J.
Personal
Communication
D. Heteroploidy 20: 2 (1979). A.
(1978).
in IUD-associated
Oral and intrauterine
Am J Obstet Gynecol 132:
in
chromosomiques 641-646 (1970).
pregnancy
(Editorial).
contraception: a 1978 risk 92-106 (1978).
8.
Tietze, C. Evaluation of intrauterine devices. Ninth Progress Report, Cooperative Statistical Program, Studies in Family Planning No. 55 (1970).
9.
Chang, C.C. and Tatum, J.H. Absence of teratogenicity of intrauterine copper wire in rats, hamsters and rabbits. Contraception 7: 413-434 (1973).
10.
Tamaya, T., Nakata, Y., Ohno, Y., Nioka, S., Furuta, M. and Okada, H. The mechanism of action of the copper IUD. Fertil Steril 27: 767-772 (1976),
11.
Parr, E.L. and Shirley, R.L. Embryotoxicity of leukocyte extracts and its relationship to intrauterine contraception. Fertil Steril 27: 1067-1077 (1976).
12.
Cur:, S., Boug, J.G. and Bou&, A. Growth characteristics of human embryonic cell lines with chromosomal anomalies. Biomedicine 21: 233-236 (1974).
13.
Jonasson, J., Therkelsen, A.J., Lauritsen, J.G. and Lindsten, J. Origin of triploidy in human abortuses. Hereditas 71: 168-171 (1972).
14.
Bomsel-Helmreich, 0. Aging of gametes, heteroploidy death. Int J Gynecol Obstet 14: 98-104 (1976).
JANUARY
1980 VOL. 21 NO. 1
and embryonic
51
CONTRACEPTION
15.
Lauritsen, in Trisomy
16.
Tredway, C.R., Umezaki, C.U., Mishell, D.R., Jr. and Settlage, Effect of intrauterine devices on sperm transport in the human being. Am J Obstet Gynecol 123: 734-735 (1975).
17.
Hunter, R.H.F. and L'Englise, P.C. Polyspermic fertilization following tubal surgery in pigs with special reference to the role of the isthmus. J Reprod Fertil 24: 233-246 (1971).
18.
In vitro Hurst, P.R., Jefferies, K., Dawson, K.J. and Eckstein, P. -development of preimplantation embryos recovered from IUD-bearing J Reprod Fertil 54: 413-417 (1978). mice.
19.
Roy, S., Klein, T.A., Scott, J.Z., Rletsky, Diagnosis of pregnancy with a radioreceptor Gynecol 50: 401-406 (1977).
20.
A simple and sensitive nonradioactive Banik, U.K. and Givner, M.L. method for the detection of urinary human chorionic gonadotropin and diagnosis of early human pregnancy. I. Multiple-unit test. Fertil Steril 32: 420-425 (1979).
21.
Banik, U.K., Hirsch, M.A., Irvine, D.S., Krupey, J., Hurwitz, A., Singh, K., Wetzel, .I., and Givner, M.L. A simple and sensitive nonradioactive method for the detection of urinary human chorionic gonadotrophin and diagnosis of early human pregnancy. II. Singleunit test. Fertil Steril 32: 426-432 (1979).
52
J.B. and Friedrich, U. Origin of the extra chromosome 16. Clin Gen 10: 156-160 (1976). C.S.
O.A. and Mishell, D.R., Jr. assay for hCG. Obstet
JANUARY
1980 VOL. 21 NO. 1