A Timeline of Pregnancy Testing
One of the earliest written records of a urine-based pregnancy
test can be found in an ancient Egyptian document. A papyrus
described a test in which a woman who might be pregnant could
urinate on wheat and barley seeds over the course of several
days: “If the barley grows, it means a male child. If
the wheat grows, it means a female child. If both do not grow,
she will not bear at all.” Testing of this theory in 1963
found that 70 percent of the time, the urine of pregnant women
did promote growth, while the urine of non-pregnant women and
men did not. Scholars have identified this as perhaps the first
test to detect a unique substance in the urine of pregnant women,
and have speculated that elevated levels of estrogens in pregnant
women’s urine may have been the key to its success.
Middle Ages through the Seventeenth Century
Using visual aspects of urine to detect pregnancy became a popular
method. In Europe, so-called “piss prophets” claimed
to be able to diagnose many different conditions and diseases
by the color of urine. In a 1552 text, pregnancy urine was described
as: “clear pale lemon color leaning toward off-white,
having a cloud on its surface.” Other tests included mixing
wine with urine and observing the results. Indeed, alcohol reacts
with certain proteins in urine, so this may have had a moderate
Various theories abounded, such as the possibility that pregnancy
urine contained certain identifiable crystals or bacteria. Scientists
did not know enough about pregnancy to develop a reliable test.
However, for sexually active women, the best method for diagnosing
pregnancy remained careful observation of their own physical
signs and symptoms (such as morning sickness).
Many physicians began to describe the workings of chemicals
in the body, suggesting that “internal secretions”
by certain organs were crucial to an understanding of human
biology. Ernest Starling named these chemical messengers “hormones.”
American public health advocates started to encourage women
to see their doctors as soon as possible after pregnancy was
suspected. Prenatal care was found to improve the health of
both infants and mothers, even though most women would not see
a doctor or midwife until well into the pregnancy.
Research on human reproduction intensified in the early twentieth
century. Ludwig Fraenkel described the corpus luteum, the glandular
mass that forms in women’s bodies during the normal menstrual
cycle that we now know is supported by hCG during pregnancy.
He identified some hormones that had a role in female reproduction,
naming the hormone that promoted gestation, progesterone. Progesterone
was isolated (an important step in the study of hormones) in
Independently, scientists in several laboratories across Europe
described the presence of a substance that promotes ovary development
and growth in rabbits and mice. In Germany, Selmar Aschheim
and Bernhard Zondek noted that this substance specifically affected
the formation of the corpus luteum.
Scientists recognized that there is a specific hormone (now
known as human chorionic gonadotropin (hCG)) that is only found
in pregnant women.
Aschheim and Zondek described a test (known as the A-Z test)
which identified the presence of hCG in urine. To test for pregnancy,
a woman’s urine was injected into an immature rat or mouse.
If the subject was not pregnant, there would be no reaction.
In the case of pregnancy, the rat would show an estrous reaction
(be in heat) despite its immaturity. This test implied that
during pregnancy there was an increased production of the hormone.
During early studies of the A-Z test, the scientists discovered
that testicular tumors could produce hCG as well.
Hormone research blossomed in this period. Scientists in several
different laboratories developed bioassays (special tests using
animals or live tissue) to identify hCG by injecting samples
to induce ovulation in rabbits, frogs, toads, and rats. These
tests were expensive, required the sacrifice of several animals,
and slow, often taking days to get results. The tests were also
insensitive when measuring hormone levels to diagnose pregnancy
because of the similarity between hCG and another substance,
luteinizing hormone (LH). Most bioassays were in fact unable
to distinguish between the two except at extraordinarily high
rates of hCG.
Herbert Evans discovered that when injected with certain fluids
from the female glands a female rat would grow an abnormally
large corpus luteum. These fluids were hormones now known as
In the next few decades laboratory scientists increased their
level of interest in the study of human reproduction and on
the role of ovaries and testes in human development.
The First International Conference of Standardization of Sex
Hormones was held in London, marking the culmination of a decade
of increased interest in the chemical properties of sex hormones
rather than the previously limited focus on biological function.
Popular childbirth books began to encourage women to visit a
doctor’s office for confirmation of pregnancy rather than
relying on “old wives’ tales” for the diagnosis.
Gonadotropins were first extracted from human pituitary glands.
A “hemagglutination inhibition test” for pregnancy
was developed by L. Wide and C.A. Gemzell. Because it used cells
in the testing process, this test was an immunoassay rather
than a bioassay. The test used purified hCG, mixed with a urine
sample and antibodies directed against hCG. In a positive pregnancy
test, the red cells clumped, displaying a particular pattern.
This test was much faster and cheaper than the old bioassay,
but still relatively insensitive, especially for early diagnosis
of pregnancy. The test also cross-reacted with various medications.
JV: The problem with these kinds of tests is that
there can be some substances in the urine to give a false-negative
or a false-positive test at a pretty high frequency, so you
had to be careful. It was important to identify what were the
interfering substances that gave false results.
Important disease research in this period led to more knowledge
about how hormones, steroids, and antibodies work in the human
body. In the next decade, NICHD scientists would transfer these
principles to their studies of reproductive hormones such as
JV: The first principle of developing a radioimmunoassay
was the result of a person making the observation that, when
patients with diabetes mellitus were treated with insulin, they
developed a circulating antibody. Then after that, we started
inducing antibody in animal models, and the rest is history.
JV: There were very few places that were doing reproductive
endocrinology research [in the late 1960s and early 1970s] because
they didn’t have purified hormones, and there were just
some very tedious ways of doing things. To compare the research
tools we had back in the late 1970s to now, it’s like
Neanderthal to modern man. [There is] no comparison. It took
brute force to get some things done.
A. R. Midgley described the first radioimmunoassay for hCG,
but the test still could not differentiate between hCG and luteinizing
hormone. Several other laboratories reported improvements on
this test, but did not solve this basic problem.
Two things came together in this period along with the so-called
sexual revolution: increased research on reproductive health
and a heightened desire (brought on by both improved prenatal
care and legal abortion) to detect pregnancy as early as possible.
Beginning in the 1970s, prenatal care and prenatal testing became
more routine in the American health care system.
"A preliminary screening test for
pregnancy," courtesy of the Food and Drug Administration
Tests available to doctors and technicians included Wampole’s
two-hour pregnancy test. The test could be done as early as
four days after a missed period. In the packaging materials,
the man pictured performing the test wore a laboratory coat,
indicating that it was not intended for home use. Besides the
equipment in the kit, (two test tubes, a plastic rack, a bottle
of “control solution,” a bottle of “hCG-antiserum”
and a bottle of “cell suspension”), testers would
need a small funnel and filter paper or centrifuge, clean pipettes
or syringes, and saline solution in addition to a urine sample.
Scientists at NIH learned more about the properties of hCG.
They were specifically interested in which parts of the hormone
showed biologic activity. Using various methods, they identified
two subunits of hCG and focused on the beta-subunit. They found
that the beta-subunit is where the immunologic and biologic
specificity of hCG resides (what makes it different from other
hormones). Using animal models, they took advantage of this
discovery to develop a specific antiserum for measuring the
hormone in humans.
In a 1972 textbook on gonadotropins, Vaitukaitis and Ross noted
that: “Common antigenic determinants [biological characteristics]
among hCG, LH, FSH, and TSH have made the production of specific
antisera for radioimmunoassay difficult.” However, the
team was close: “the recent isolation and separation of
subunits…have provided unique materials with which these
questions could be explored.”
Vaitukaitis, Braunstein, and Ross published their paper
describing the hCG beta-subunit radioimmunoassay that could
finally distinguish between hCG and LH, therefore making it
potentially useful as an early test for pregnancy. See Vaitukaitis,
J.L., Braunstein, G.D., and Ross, G.T. (1972) “A radioimmunoassay
which specifically measures human chorionic gonadotropin in
the presence of human luteinizing hormone.” American
Journal of Obstetrics and Gynecology, 113, 751-8.
The first edition of Our Bodies, Ourselves, the women’s
health manual written by the Boston Women’s Health Collective,
noted that available pregnancy tests were most accurate if done
two weeks after the missed period. Though the authors insisted
that instructions for “collecting and submitting your
urine are simple,” modern readers might disagree. “Drink
no liquids after dinner the night before,” the text instructed,
“then as soon as you awake in the morning collect a urine
sample in a clean, dry, soap-free jar and take it to a laboratory.”
Another possibility was sending the urine sample to a laboratory
in North Carolina, after first writing to request the test kit.
Though the test was not yet widely available, NIH scientists
spread the word about the new radioimmunoassay. At first, the
test was found most useful for clinicians in testing and following
patients being treated for hCG-secreting tumors. The sensitive
radioimmunoassay could tell the doctors if the chemotherapy
treatments had worked.
JV: We were doing assays for people all over the place.
We felt ethically that we had to because it wasn’t available
anyplace else. So we used to give out a lot of antiserums to
research labs and show them how to set up the assays.
From Directions and Technical Information
on UCG-TEST, 1970, courtesy of Special Collections, Northwestern
FDA approval was sought by Warner-Chilcott for e.p.t, the “Early
Pregnancy Test” later known as the “Error Proof
Test.” e.p.t would become the first home pregnancy test
kit on the market in the United States. The makers of e.p.t
worked with the FDA to meet all the requirements of the 1976
Medical Devices Act. (The new regulations divided medical devices
into three classes depending on potential for harm and misuse.)
Approval was also granted to three other tests that were deemed
“Substantially equivalent:” Predictor, ACU-TEST,
Several articles in the American Journal of Public Health
stated that public health would be better served if the average
consumer could purchase a home pregnancy test and use it reliably
in her own home.
By the end of 1977, e.p.t was ready for the American market.
(Because of requirements for the specific wording on packaging
and other last-minute details, there is a lag time between FDA
approval and wide availability of most medical devices.) In
a “Dear Pharmacist” letter from Warner/Chilcott,
drug stores were informed that “the e.p.t consumer advertising
campaign has been designed to direct the consumer to their drug
store to purchase e.p.t”
e.p.t was advertised in major women’s magazines including:
Mademoiselle, McCall’s, Redbook,
Family Circle, Ladies’ Home Journal, Good
Housekeeping, and Vogue. Advertisements appeared
later in the year for Predictor, Answer, and ACU-TEST.
The e.p.t test of 1978 was described to the public in Mademoiselle:
“For your $10,” the article notes, “you get
pre-measured ingredients consisting of a vial of purified water,
a test tube containing, among other things, sheep red blood
cells…as well as a medicine dropper and clear plastic
support for the test tube, with an angled mirror at the bottom.”
The test took two hours, and was more accurate for positive
results (97%) than for negative (80%). Advantages, noted Mademoiselle,
included “privacy and not having to wait several more
weeks for a doctor’s confirmation, which gives you a chance,
if pregnant, to start taking care of yourself…or to consider
the possibility of early abortion.” (Mademoiselle,
April 1978, p. 86)
McCall’s magazine claimed that “physicians
we interviewed about the tests endorse the concept.” But
the editors cautioned that women who get negative results and
who still suspect pregnancy should not wait ten days to take
the test again “but should seek medical help as soon as
possible.” (McCall’s, March 1978, p. 46)
Taking the test at home, noted a 1979 article in Family Planning
Perspectives, both protected the privacy of a woman who
might not want her doctor to know she is sexually active and
gave women a new opportunity to take an active role in their
own health care.
Research increased and educational campaigns were launched to
identify the importance of folic acid in early pregnancy and
to warn of the dangers of various environmental hazards and
alcohol to a developing fetus.
Advances in the technology of pregnancy tests included the development
of new types of antibodies and the use of enzyme labels in place
of radioactive labels.
The next generation of home pregnancy tests was ushered in with
FDA approval of Clearblue Easy’s digital pregnancy test.
In place of a thin blue line, the indicator screen will now
display either “pregnant” or “not pregnant.”
JV: The home pregnancy test is probably the most widely
used test besides the hematocrit and hemoglobin [the blood test
to measure red blood cells and iron levels that is part of the
blood workup done regularly at doctors’ offices].
Note: Quotations labeled “JV”
are from an interview with Judith Vaitukaitis, August 18, 2003.
Quotations labeled “GB” are from a telephone
conversation with Glenn Braunstein, October 3, 2003.
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Advertisement for ACU-TEST, American
Journal of Public Health, January 1979.
"Dear Pharmacist" letter from Warner-Chilcott,
c. 1977. Courtesy of the National Museum of American History,