Human Reproduction Vol.21, No.7 pp. 1645–1650, 2006
doi:10.1093/humrep/del067
Advance Access publication April 10, 2006.
OPINION
A.R.T. and history, 1678–1978
Gary N.Clarke
Andrology Unit, The Royal Women’s Hospital, Carlton, Victoria, Australia
To whom correspondence should be addressed at: Andrology Unit, The Royal Women’s Hospital, Carlton, Victoria 3053, Australia.
E-mail: [email protected]
Louise Brown, the first baby conceived after IVF, was born on 25 July 1978 and turned 27 last year. From one perspective, her birth can be seen as the culmination of 300 years of medical and scientific investigation aimed at understanding
the fascinating process of reproduction. This essay was written as a tribute to mark the unique contribution to assisted
reproductive technology (ART) which resulted from the collaboration of a scientist, Bob Edwards, and a clinician,
Patrick Steptoe, who pioneered the successful clinical use of IVF. This article was not intended to be a conventional history of science, but instead has primarily focused on those early discoveries which in the author’s opinion were critical
to our current understanding of mammalian reproduction. There are some digressions and many omissions necessitated
by attempting to cover 300 years in a relatively short essay. In particular, there is no mention of endocrinology because
this area has been covered in numerous reviews and books. The main sources of historical information for this article
were the authoritative books of Professor Cole (1930), Dr Elizabeth Gasking (1967), Professor John Farley (1982),
Dr Fielding H. Garrison (1929) and the Philosophical Transactions of The Royal Society or Letters collated from the latter.
Introduction
The process of reproduction has long fascinated mankind, particularly from philosophical and scientific perspectives. For
example, Aristotle devoted a considerable amount of his time
to study the process of reproduction, and William Harvey
(1578–1657) performed exacting trials on the King’s herd of
deer and wrote a major treatise on the subject (‘De generatione
animalium’, 1651) which contains in the frontispiece a drawing
of Jove releasing a plethora of creatures from an egg on which
is inscribed his famous dictum:
Ex ovo omnia
The meaning of Harvey’s dictum was expressed poetically by
his friend Martin Lluelyn:
That both the hen and housewife are so matcht,
That her son born, is only her son hatcht,
That when her teeming hopes have prosp’rous been,
Yet to conceive, is but to lay within.
Harvey’s work is often regarded by historians of science as
defining the boundary between the early era which relied heavily
on folklore, superstition and casual observation, and the recent
period during which experimentation and more exacting observations of nature became the modus operandi for many scientists
and natural philosophers. However, although it was obvious to
most people that many creatures were hatched from eggs and that
the male was essential for sexual reproduction, the discovery and
experimental study of spermatozoa and mammalian oocytes
required a new scientific instrument, albeit initially a very crude
one. One of the first microscopists was Antonj van Leeuwenhoek
(1632–1723) who, amongst his many other discoveries, was the
first to conduct rigorous observations on human spermatozoa.
The discovery of spermatozoa and the doctrine
of preformation
They are formed of a transparent substance, their movements are
very brisk and their shape is similar to that of frogs before their
limbs are formed. This discovery, which was made in Holland
for the first time, seems very important, and should give employment to those interested in the generation of animals.
This is an extract from a letter written by Christiaan Huygens
in 1678 and published in Paris in the Journal des Savants (see
Gasking, 1967). It was the first published description of spermatozoa following van Leeuwenhoek’s original observations
of the microscopic tadpole-like creatures in human semen
which were known at the time as ‘animalcules’ or ‘spermatick
worms’. Antonj van Leeuwenhoek had previously conveyed
his observations in a letter to the Royal Society in 1677, which
was eventually published in the Philosophical Transactions in
1679 (Leeuwenhoek, 1941). The Fellows were very interested
in the ‘living creatures’ in semen and consequently the Secretary wrote back to van Leeuwenhoek with a request that he
examine the semen of other species. There was a subsequent
dispute over priority with Nicolas Hartsoeker, who had also
reported his observations on rooster semen later in 1678, subsequently claiming that he had first discovered the animalcules
© The Author 2006. Published by Oxford University Press on behalf of the European Society of Human Reproduction and Embryology. All rights reserved.
For Permissions, please email: [email protected]
1645
G.N.Clarke
(see Gasking, 1967). There is strong circumstantial evidence,
though, that both Huygens and Hartsoeker had seen Leeuwenhoek’s
letter prior to publishing their own letters (Cole, 1930). However,
it was Johan Ham, a medical student at Leiden whose observations on spermatozoa apparently antedated those of both van
Leeuwenhoek and Hartsoeker, but although van Leeuwenhoek
referred to Ham in his letter to The Royal Society, the young student never published his own observations (Figures 1 and 2).
The discovery of the vast numbers of motile creatures in
semen stimulated a lot of debate as to their origin and function
because at the time it was far from obvious that they might be
the agents of conception. Many scientists were convinced that
they must be parasites; some speculated that they were essential for conception, whilst others thought that they were a normal component of semen, but that they performed a secondary
role such as mixing the other seminal components. The arguments
about the role of spermatozoa may be somewhat astonishing
Figure 1. Microscope made by van Leeuwenhoek in 1673.
when viewed retrospectively, but it is important to remember
that the discovery of the mammalian oocyte was not reported
until 1827 by Carl Ernst von Baer and that Edouard van
Beneden first observed fertilization in mammals in 1875. Consequently, many scientists up until around 1850 continued to
regard spermatozoa as parasites (the name spermatozoa was
first used by von Baer in 1827 and obviously reflects this
view). Thus, in 1835 the anatomist Richard Owen classified
spermatozoa as parasites and relegated them to the Entozoa
(order Prothelmintha). Subsequently, the investigations of the
Swiss scientist, Albert von Kolliker (1841) were pivotal in
demonstrating that spermatozoa were not parasites, but motile
autologous cells which developed from testicular cells by the
process of histogenesis. In his words, sperm were:
organized parts of the seminal fluid, elementary parts analogous
to the blood corpuscles
Even though the exact role of the spermatozoa in generation
was a controversial topic which was vigorously debated for
many years, their discovery did open a new front in the ongoing philosophical/scientific debate over preformation (the doctrine that the complete, preformed embryo was present in the
generative organs of either the male or the female parent),
which was often coupled with emboitement (the doctrine that
the complete minature organism has always been encapsulated
within its own parent, analogous to the Russian Babushka dolls
which are made progressively smaller so they fit one inside the
other). The ovists argued that the preformed germ or embryo
came from the female, whilst their protagonists asserted that it
came from the seminal contribution. On the basis of his recent
discovery of spermatozoa or animalcules as they were then
known, in 1683 van Leeuwenhoek proposed the animalculist
version of the preformation theory which contended that the
spermatozoa contributed the essential germ to be nurtured by
the female. Although fascinating historically, particularly from
a philosophical standpoint (see Wilson, 1995; for in-depth discussion), from our current perspective, this was a pointless
debate, so we can shortly return to some of the crucial experimental work which ultimately clarified the role of spermatozoa
and oocytes in the process of generation or fecundation. Before
doing so, it is worth digressing to discuss the often quoted and
misquoted homunculus story.
The homonculus
According to Cole (1930), the first author to publish a representation of a preformed fetus in the sperm head was Hartsoeker
(1694) in his Essay de Dioptrique. However, Hartsoeker did
not pretend that he had actually seen the homunculus, in the
text he stated:
if we could see the little animal through the skin which hides it
we might possibly see it as it is represented in the illustration
Figure 2. Van Leeuwenhoek’s drawings of spermatozoa.
1646
The homunculus gained credibility with the report by Dalenpatius
(1699), wherein he stated explicitly that he had observed a
minute human form inside the sperm head. Unusually, his letter was published simultaneously in Amsterdam, London and
Edinburgh, which Cole (1930) considered was indicative of it
A.R.T. and history, 1678–1978
being ‘an organized and serious attempt to deceive the public’.
The real identity of Dalenpatius was eventually revealed by
Jean Astruc in 1740 in his book on venereal diseases. Astruc
had been a friend of a man by the name of Plantade who was
‘given to jesting’ and had composed the letter in order to
amuse himself, and that the assumed name was an anagram of
Plantadeius which was the latinized form of his real name (see
Cole, 1930). Unfortunately, many bona fide natural philosophers believed his report of the homunculus, which was then
discussed seriously in various publications. Thus the homunculus acquired a degree of currency in the literature before the
hoax was eventually uncovered. In 1749 George-Louis de Buffon referred to Dalenpatius observations as:
repugnant to the repeated experience and observation of all those
who have hitherto made enquiries into this subject.
It is quite possible that Plantade was satirizing the beliefs held
by some of the preformationists in the animalculist school in an
attempt to ridicule and discredit them (Cole, 1930). Whatever
his intentions, however, once unleashed the homunculus had
various reincarnations over the next 150 years. Thus, as late as
1750, possibly as another hoax, Gautier d’Agoty reported and
figured a mal-proportioned fetus in human semen (see Cole,
1930) (Figure 3).
Figure 3. A Dutch draper with a passion for the microscopic study of
nature, van Leeuwenhoek made around 247 microscopes with which
he made many exciting discoveries which he communicated to The
Royal Society in some 375 letters during his long life. He was elected
a Fellow of The Royal Society (FRS) in 1680. He also sent 27 letters
to the French Academy of Sciences. He was the first to describe spermatozoa, the striped appearance of voluntary muscle, the structure of
the crystalline lens, and he also observed the capillaries linking the
veins and arteries, thus completing what Harvey had started. The portrait shown and the biographical information above were derived from
Garrison (1929). (Visit Brian Ford’s website www.sciences.
demon.co.uk/∼bjford.htm for a wealth of information on Leeuwenhoek,
his original specimens sent to The Royal Society and the performance
of his simple microscopes.)
Elucidation of the role of spermatozoa in fertilization
The Italian scientist Lazaro Spallanzani (1729–1799) was a
very good experimentalist who turned his attention to semen in
the 1770s. He examined semen from mammals, fish and
amphibians and confirmed the presence of spermatozoa. In
experiments with frogs, he was able to show that oocytes
would only develop into tadpoles after contact with semen,
possibly the first example of IVF. He also successfully artificially inseminated a spaniel bitch of which he wrote:
The success of this experiment gave me more pleasure than I
have ever felt in any of my other scientific researches.
In subsequent experiments, Spallanzani found that the fertilizing
ability of semen was destroyed by dessication or heat or by
mixing it with certain amounts of vinegar, salt or spirits. He
also observed that filtration of semen could remove the fertilizing capacity, but that if the material collected onto the filter
was immediately redissolved in water, then fertilization could
occur. Unfortunately, because of the prevailing dogma of ovist
preformation, the results of Spallanzani’s elegant experiments
were misinterpreted both by himself and by the wider scientific
community (see Gasking, 1967). In essence, because Spallanzani
believed that spermatozoa were parasites, he assumed that
some other component of the seminal fluid which had also
been retained on the filters must be the fecundating agent.
In the early part of the nineteenth century, the Swiss physician, Jean-Louis Prevost and the French scientist, Jean-Baptiste
Dumas made a major contribution to the field with their
exhaustive comparative investigations showing that spermatozoa were present in the testes of many different animals (see
Gasking, 1967). They concluded that the spermatozoa must be
produced in the testes. Their work was heavily criticized at the
time because it was perceived as a reversion to the outdated
animalculist theories of the eighteenth century, but in the
longer term it lent significant weight to the mounting evidence
for the importance of the spermatozoal contribution to the
reproductive process. Most of their research work was published in the Annales des Sciences Naturelles in the period
1821–30. Contrary to the generally accepted view, they
asserted that spermatozoa were not parasites but instead were
the ‘result of a true secretory action and the active principle of
fecundation’. The reason for the conflict with most of their scientific peers was summarized by Farley (1982):
only the animalculists and pollenists of the seventeenth and
eighteenth centuries believed that the sperm and pollen grains
played any role in generation. According to them, the sperm and
pollen contained a preformed embryo which needed to be
implanted inside the egg in order for development to begin.
These theories were viewed as historical relics by naturalists of
the early nineteenth century. By then fecundation was seen as
either the dynamic interaction of male and female fluids or the
interaction of a male fluid with a preformed egg. Both theories
explicitly denied that spermatic animalcules or pollen particles
had any role to play in the reproductive process.
In further experiments Prevost and Dumas demonstrated convincingly that motile spermatozoa were essential for the fertilization of frog oocytes. Their working hypothesis was that each
1647
G.N.Clarke
spermatozoon fertilized a single oocyte. They accumulated significant evidence to support their hypothesis in a series of experiments wherein they added a known number of spermatozoa to a
known, but always greater number of oocytes. They observed
that the number of oocytes which underwent development was
always lower than the number of spermatozoa added.
Prevost and Dumas also repeated Spallanzani’s filtration
experiments, but because they were open to the possibility that
the spermatozoa were the fecundating agent, they concluded
that filtration prevented fertilization by removing them from
the seminal fluid.
Final confirmation of the necessity for sperm–oocyte fusion
during sexual reproduction was the result of key observations
in a number of species over several decades (see Farley, 1982).
As early as 1852, Henry Nelson had reported observing that
the sperm of Ascaris entered the completely transparent ovule,
and he claimed that his investigation was:
the first in which the act of the penetration of the spermatozoa
into the ovum has been distinctly seen and clearly established.
George Newport (1853) subsequently reported seeing fertilization in amphibians, and then Edouard van Beneden (1875) and
Oscar Hertwig (1876) reported their respective observations of
mammalian fertilization. Herman Fol, who, like Hertwig, had
trained under Ernst Haeckel at Jena, reported his observations
on fertilization in starfish in 1879. He described the penetration
of a single sperm into the oocyte to form the male pronucleus,
which moved within the oocyte cytoplasm to fuse with the
female pronucleus. In addition, van Beneden (1879) described
the reduction in chromosome number in the gametes of Ascaris
and then proposed that the chromosome must be the physical
vehicle of inheritance (1883).
corpus luteum. In 1794, William Hunter noted that there were
two corpora lutea associated with several cases of twins which
he investigated versus only one with a number of singleton
pregnancies (Short, 1977).
During the early nineteenth century, Prevost and Dumas
made vital contributions in this area with their work on rabbits
and dogs during which they carefully observed the ovaries both
before and after coitus. Having noted significant alterations in
the Graafian follicles and the formation of the corpus luteum, a
few days later they observed tiny ovoid bodies in the oviducts.
They subsequently speculated that mammalian oocytes were
probably fertilized in the oviducts.
Another very important contributor to our knowledge of the
reproductive process was Carl Ernst von Baer, an embryologist
par excellence. In 1826 he identified the mammalian oocyte
during his studies on the ovary of a bitch. This discovery was
reported in his classic monograph of 1827, ‘De ovi mammalium et homonis genesi’ published in Leipzig (see Poynter,
1968). The descriptions of his work were both lucid and prescriptive so that fellow scientists could readily confirm his
observations. In exhaustive comparative studies, he catalogued
the early stages of embryonic development and in so doing he
laid solid foundations for later investigators to integrate them
with the cell theory of Mathias Schleiden and Theodor
Schwann (1838) (Figure 4).
Artificial insemination
Because the subject was shrouded in secrecy, it is impossible
to accurately document when artificial insemination was first
The discovery of the mammalian oocyte and early
embryological studies
According to Short (1977), Soranus of Ephesus (ca. 50 AD)
was the first anatomist to publish a detailed description of ovaries or didymi (paired organs), as he called them. The first
authoritative description of ovarian follicles and the corpus
luteum was made by Vesalius in 1555, whilst his student Fallopius
described the Fallopian tubes in 1562 and made further observations on the ovaries (Short, 1977). Fabricius (1533–1619)
studied under Fallopius and subsequently became a great anatomist who published a superbly illustrated book (‘De Formato
Foetu’, 1604) on the comparative anatomy of the female reproductive tract. Fabricius in turn was the teacher of William
Harvey (1578–1657) who published the results of his studies
on reproduction in 1651 in ‘De Generatione Animalium’.
Although Harvey’s motto was ‘ex ovo omnia’, as mentioned in
the ‘Introduction’, he wrongly believed that the oocyte was the
product of conception. He was also unable to find any evidence
that the ovaries played a significant role in reproduction (Short,
1977). In 1667, Niels Stensen proposed that the ‘female testes’
contained oocytes and were analogous to the ovaries of
oviparous animals. Another significant contributor was the
Dutchman, Regnier de Graaf after whom the Graafian follicles
are named, but who also gave the first detailed account of the
1648
Figure 4. The father of modern embryology who developed the
germ-layer theory and made exacting studies of organogenesis and
morphogenesis. Considering the relatively crude microscopes and the
fact that he did not have a microtome, the results obtained by von Baer
were astounding. He reported his discovery of the mammalian oocyte
in 1827 and described the notochord. This great scientist conducted
extensive investigations into comparative anatomy which culminated
in his classification of animals into four main groups—Radiata, Mollusca, Articulata and Vertebrata. The portrait shown and the biographical information above were derived from Garrison (1929).
A.R.T. and history, 1678–1978
used by medical practioners. Dr John Hunter has often been
cited as having performed the first documented artificial
insemination in about 1790. However, in an article by Poynter
(1968) on the history of artificial insemination, it was stated
that Hunter’s case was published after his death by his executor
Sir Everard Home in 1799. Apparently, in about 1776, a linen
draper suffering from infertility associated with hypospadias
became a patient of Dr Hunter’s. After instructions from the
doctor, he successfully inseminated his wife at home using a
syringe.
The first published account of a successful human artificial
insemination actually performed by a doctor concerned a treatment performed on 5 June 1838 in France by Dr Girault. His
patient, a young countess, gave birth to a normal son on 1
March 1839. This case was the first in a series of 12, which
were recorded in a paper published in 1868 (L’Abeille Medicale, volume 25, pages 409–17; cited in Poynter, 1968). In
1866, the renowned American gynaecologist Dr James Marion
Sims (1813–1883) published his book on sterility which
included a chapter on artificial insemination. This book stimulated an extremely vigorous debate around the medical, moral
and ethical issues surrounding artificial insemination. As stated
by Poynter ‘The effect of the publication of Sim’s book in
France was to stimulate a more open discussion in medical circles where it had been practiced with discretion for years’. The
practice of artificial insemination gained acceptance quite rapidly in France and by 1888 a book by Dr Lajatre from Bordeaux was in its 10th edition (see Poynter, 1968). The first
published reference to donor insemination was made by an
Italian Paolo Mantegazza in 1887.
With the exception of France, the wide acceptance of artificial insemination amongst the medical establishment took
many decades. It was still extremely controversial in the USA
in the 1940s when Dr Clair E. Folsome (1943) published his
Critical Review, and donor insemination was still quite controversial in Australia in the 1970s when the first sperm banks and
public hospital-based donor insemination programmes were
established.
Cryopreservation of reproductive tissues
It is likely that the first investigations of the effects of low
temperatures on spermatozoa were carried out by Lazaro
Spallanzani (1776), who subjected them to:
the freezing cold of winter and it’s snow
The first to discuss the possible uses of sperm banks was Paolo
Mantegazza, the Professor of Pathology at Pavia, who wrote in
1866:
It might even be that a husband who has died on a battle-field can
fecundate his own wife after he has been reduced to a corpse and
produce legitimate children after his death.
However, this did not become a realistic proposition until the
discovery of the very effective cryoprotective properties of
glycerol by Polge et al. (1949). The first human births resulting
from artificial insemination of cryopreserved semen were reported
by Bunge and Sherman in 1953. By 1976 approximately 1500
births had resulted from this procedure (Sherman, 1976), and
initial concerns about possible genetic damage caused by the
cryopreservation process were abating. Whittingham et al.
(1972) reported the successful cryopreservation of mouse
embryos at –196 and –269°C. The first human pregnancies
resulting from the transfer of cryopreserved embryos were
reported by Zeilmaker et al. (1984).
Towards IVF
Early attempts at IVF using animal gametes were published by
Shenk (1878) and Long (1912), but these reports have dubious
significance because relatively inexact criteria were used to
define fertilization and also because the role of temperature
changes in pathogenic activation was not identified until 1936
by Pincus and Enzmann (see Gwatkin, 1977). Perhaps 1951
could be seen as a critical boundary defining the beginning of
the modern era with the independent identification of sperm
capacitation by both Colin Austin (1914–2004) and M.C.
Chang. Subsequently, Austin and Bishop (1958) published the
first evidence for the occurrence of the acrosome reaction prior
to sperm penetration of the zona pellucida. The first report of
successful IVF using sperm-capacitated in vitro was made by
Yanagimachi and Chang (1963). These important contributions
laid the groundwork which would ultimately lead to successful
human IVF (see Bavister’s, 2002 review for detailed discussion and bibliography relating to capacitation and acrosomereaction and Blandau, 1980; Wood and Trounson, 2000 for
more detail on the early history of IVF).
The first report on human IVF was by Edwards et al. (1969)
who observed pronuclear formation in a few oocytes. In 1970
they reported that they had obtained embryonic development to
the 16-cell stage (Edwards et al., 1970), and ultimately in 1978
they reported the first birth resulting from IVF (Steptoe and
Edwards, 1978), with more detail presented in a paper delivered at the Scientific meeting of The Royal College of Obstetricians and Gynaecologists on 26 January 1979, entitled
‘Pregnancies following implantation of human embryos grown
in culture’, followed by further publications in 1980
(Edwards et al., 1980; Steptoe et al., 1980). Their reports
were followed closely by reports of successful IVF pregnancies in Melbourne (Lopata et al., 1980; Trounson et al., 1981).
Postscript
The enormous impact of IVF in treating infertility can be
gauged by examination of the accurate data compiled by the
National Perinatal Statistics Unit for Australia and New Zealand
over the first two decades after its initial introduction. Thus, in
1979 there were two IVF pregnancies which grew to 1905 in
1989 and 4952 in 1999 with a total of 40 585 IVF pregnancies
during that period (Hurst and Lancaster, 2001).
An indication of the global impact can be obtained from the
‘World collaborative report on assisted reproductive technology, 1998’ which was presented at the 17th World Congress
on Fertility and Sterility held in Melbourne, Australia in 2001
and subsequently published in the Proceedings of this meeting
(Adamson et al., 2002). The global data is incomplete but were
1649
G.N.Clarke
collected from 1504 clinics operating in 44 countries, who performed a total of 390 000 treatment cycles resulting in 84 594
children born from conceptions in 1998 alone. This was compared with an estimated 45 000 children born from conceptions
initiated in 1995. We can extrapolate conservatively from this
data to estimate that over the last 25 years between 250 000 and
500 000 children have been conceived through IVF procedures.
References
Adamson D, Lancaster P, de Mouzon J, Nygren KG and Zegers-Hochschild F
(2002) World collaborative report on assisted reproductive technology,
1998. In Healy DL, Kovacs GT, McLachlan R and Rodriguez-Armas O
(eds) Reproductive Medicine in the Twenty-First Century. Parthenon
Publishing, Boca Raton, pp. 209–219.
Austin CR (1951) Observations on the penetration of sperm into the mammalian egg. Aust J Sci Res (B) 4,581–592.
Austin CR and Bishop MWH (1958) Some features of the acrosome and perforatorium in mammalian spermatozoa. Proc R Soc Med 67,925–927.
Bavister BD (2002) Early history of in vitro fertilization. Reproduction
124,181–196.
Blandau RJ (1980) In vitro fertilization and embryo transfer. Fertil Steril 33,3–11.
Chang MC (1951) The fertilizing capacity of spermatozoa deposited into the
fallopian tubes. Nature 168,697–698.
Cole FJ (1930) Early Theories of Sexual Generation. Oxford University Press,
Oxford.
Edwards RG, Bavister BD and Steptoe PC (1969) Early stages of fertilization
in vitro of human eggs matured in vitro. Nature 221,632–635.
Edwards RG, Steptoe PC and Purdy JM (1970) Fertilisation and cleavage in
vitro of preovulatory human oocytes. Nature 227,1307–1309.
Edwards RG, Steptoe PC and Purdy JM (1980) Establishing full-term human
pregnancies using cleaving embryos grown in vitro. Br J Obstet Gynaecol
87,737–756.
Farley J (1982) Gametes and Spores. Ideas About Sexual Reproduction, 1750–
1914. The Johns Hopkins University Press, Baltimore and London.
Folsome CE (1943) The status of artificial insemination. A critical review. Am
J Obstet Gynaecol 45,915–927.
Garrison FH (1929) History of Medicine, 4th edn. W.B. Saunders, Philadelphia
and London.
Gasking EB (1967) Investigations into Generation, 1651–1828. Hutchinson,
London.
1650
Gwatkin RBL (1977) Fertilization Mechanisms in Man and Animals. Plenum
Press, New York and London.
Hurst T and Lancaster P (2001) Assisted Conception, Australia and New Zealand
1999 and 2000. National Perinatal Statistics Unit. Australian Institute of
Health and Welfare, Sydney, Australia.
Leeuwenhoek A (1941) The Collected Letters, Volume II, 1676–1679. Edited
by a Committee of Dutch scientists. Swets and Zeitlinger Ltd, Amsterdam.
Lopata A (1980) Pregnancy following intrauterine implantation of an embryo
obtained by in vitro fertilization of a preovulatory egg. Fertil Steril 33,117–120.
Newport G (1853) On the impregnation of the ovum in amphibia. Philos Trans
R Soc Lond 143,233–290.
Polge E, Smith AU and Parkes AS (1949) Revival of spermatozoa after vitrification and dehydration at low temperatures. Nature 164,666.
Poynter FNL (1968) Hunter, Spallanzani, and the history of artificial insemination. In Stevenson LG and Multhauf RP (eds) Medicine, Science and Culture. The Johns Hopkins Press, Baltimore, MD.
Sherman JK (1976) Clinical use of frozen human semen. Transplant Proc
8,165–170.
Short RV (1977) The discovery of the ovaries. In Zuckerman S and Weir BJ (eds)
The Ovary, 2nd edn. Academic Press, New York.
Sims JM (1866) Clinical Notes on Uterine Surgery, with Special Reference to
the Management of the Sterile Condition. R. Hardwicke, London.
Steptoe PC and Edwards RG (1978) Birth after the reimplantation of a human
embryo. Lancet 2,366.
Steptoe PC, Edwards RG and Purdy JM (1980) Clinical aspects of pregnancies
established with cleaving embryos grown in vitro. Br J Obstet Gynaecol
87,757–768.
Trounson AO, Leeton JF, Wood C, Webb J and Wood J (1981) Pregnancies in
humans by fertilization in vitro. Embryo transfer in controlled ovulatory
cycles. Science 212,681–684.
Whittingham D, Leibo S and Mazur P (1972) Survival of mouse embryos frozen to −196°C and − 269°C. Science 178,411–414.
Wilson C (1995) The Invisible World. Early Modern Philosophy and the
Invention of the Microscope. Princeton University Press, Princeton, NJ.
Wood C and Trounson AO (2000) Historical perspectives of IVF. In Trounson
AO and Gardner DK (eds) Handbook of In Vitro Fertilization, 2nd edn.
CRC Press, Boca Raton, pp. 1–14.
Yanagimachi R and Chang MC (1963) Fertilisation of hamster eggs in vitro.
Nature 200,281–282.
Zeilmaker G, Alberda A, van Gent I et al. (1984) Two pregnancies following
transfer of intact frozen-thawed embryos. Fertil Steril 42,293–296.
Submitted on October 7, 2004; resubmitted on November 4, 2004; accepted on
November 19, 2004