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I. PENDAHULUAN
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a. Materi
b. Metode
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a. Hasil
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DAFTAR REFERENSI
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 MANUFACTURE AND EVALUATION OF VAGINAL SMEAR OF MICE

By:
Name : Safira Dwi Oktaviani
Student ID : B1B015002
Group : VII
Subgruop :1
Assistant : Sarah Nurul

PRACTICAL REPORT OF ANIMAL DEVELOPMENT

MINISTRY OF RESEARCH, TECHNOLOGY, AND HIGHER EDUCATION

JENDERAL SOEDIRMAN UNIVERSITY
FACULTY OF BIOLOGY
PURWOKERTO

2016
 I. INTRODUCTION
A. Background

Mammals anything in the world to have sexual activity that accompanies all his
life. The sexual activity is always changing, sometimes high and sometimes low.
Period showed that the females are experiencing high sexual activity indicated with
signs such as anxiety and screaming for males called the estrus. The term was
originally only indicate the presence of estrous periods of high sexual desire, which
is realized through the animal's behavior. Data obtained through experiment is known
that in the event of estrus also been changes are important in the animal, which is
closely associated with the time of ovulation, which usually coincide with the phase
of estrus (Nalbandov, 1976).
Vaginal smear is one method for observing cell types and proportions of each
cellfound in the vaginal smear. Results obtained from these observations may
determine the phase that is being experienced by female animals tested. This method
is based on reality that during estrus phase, vaginal epithelial cells undergo
cornification as a result of high levels of estrogen. Animals who want to know the
phase in the cycle is estrus female animals that have been mature gonad and not
pregnant. Estrous cycle is the distance between estrus one to the next
estrus. Recycling or estrus cycle consists of four phases, namely proestrus, estrus,
metestrus, and diestrus.
Vaginal smear is method used for identification phase estrus cycle being
experienced by female mamals. Each of estrus cycle had a different cell type. The
different cell type can be used as sign to determine a phase of estrus in female
individuals. The period between the estrus phase with subsequent estrus phase is
called the estrus cycle. Each animal has estrus cycles are different, there is a group of
animals monoestrus (estrus once a year), class poliestrus (estrus several times a year)
and poliestrus animals in season (estrus only during certain seasons of the year). One
of the estrous cycle, there are four phases: proestrus, estrus metestrus or postestrus,
and diestrus. Each of these phases are associated with changes in sexual activity and
structure in the ovary, uterus, and vagina. Period showed that the females are
experiencing high sexual activity indicated with signs such as anxiety and behaving
strangely towards males. The term estrus originally only indicate the presence of a
period of sexual desire is high, which is realized through the behavior of the animal,
but by obtaining data through experiments, it is known that in the event of estrus also
been changes are important in the animal, which is closely associated with the
current ovulation, which usually coincide with the phase of estrus (Hafez, 1968).
Preparations in the form of mice ( Mus musculus ) is used in the vaginal smear
lab because the animal had sex ripe and are not pregnant who have a fairly low age
of about 8 weeks, easy to swab sampling, and easily obtainable. In addition, these
mice belong to a class of mammals that have a relatively small body but has discount
reproductive organs are fairly complete. Estrus cycles experienced by the murine also
in the not too long so it is easy to do observations (Martini, 2000).

B. Porpose

The purpose of this lab is to create a vaginal swab of the mise ( Mus
muscullus ) and to identify the cell types in the preparations and determine estrus
phase of the test animals.

II. MATERIAL AND METODE

A. Material
 Tools used in this lab is a light microscope, along with the cover glass object, a
cotton bud and tissue.
The materials used in the lab vaginal smear is female mice ( Mus musculus )
mature gonad and not pregnant, 0.9% NaCl solution, a solution of 70% alcohol, 1%
methylene blue dye akuosa and water faucets with low discharge.

B. Method

The method used in this lab are as follows:

1. Mice ( Mus musculus ), which will be examined by means of hand-held right on

the palm melentangkannya temporarily clamped by the nape of his thumb and
forefinger. Tail clamped between palms and little finger.
2. The tip of a cotton bud moistened with a solution of NaCl 0.9% and then slowly
inserted into the vagina of mice as deep as 5 mm. Cotton swabsrotated clockwise
slowly two or three times to obtain a white liquid on a cotton bud.
3. The glass object to be cleaned using 70% alcohol and air dried. The tip of a
cotton bud that has been applied in the vagina is applied lengthwise on a glass
object with two or three rows spread in the same direction on a glass object.
4. spread the vagina drops of the solution of 1% methylene blue and wait about 5
minutes. Excessive dye cleaned with tap water drop by drop and then covered
with a glass lid.
5. Mixture was observed using a light microscope with a magnification of weak
and then strong magnification by taking into account the type and proportion of
cells in preparation. Then draw the shape of epithelial cells and leukocytes
appear on vaginal smears.
 III. RESULT AND DISCUSSION

A. Result

(A) (B)

Details:

Figure (A) : Microscopic Estrus Cycle Phase Estrus. Magnification 100x

Figure (B) : Schematic Estrus Cycle Phase Estrus

Figure Details :

1. Epithel cell
2. Leukocytes
 B. Discussion

Based on observations of smears of the vagina that has been done, it can be
seen that the visible cells are cells that are undergoing a phase of estrous end
approaching matestrus, it is seen in the presence of epithelial cells terkornification
and the presence of leukocytes albeit small, the size of the epithelial cells of large
and without core , The results of these observations in accordance with the statement
of Taylor (1994), that the estrus phase characterized by epithelial cells gore, have no
nucleus and looked pale. In this phase, estrogen production increases. There was a
decrease in the blood FSH and LH increase.
At this time of observation in mice can said, in a period of estrus estus yet
final and approached metestrus. because at this time in the observation found that
terkornifikasi epithelial tissue and also leukocytes in small quantities. According
Muchtarromah (2006) estrus phase of the final mark with cornification epithelial
cells, leukocytes or nucleated epithelial cells, epithelial cell cornification, and
leukocytes. whereas metestrus phase is the phase that is characterized by the
presence of epithelial cells and leukocytes many gore. The duration of this phase is
approximately 8 hours (Billet and Wild, 1975)
Vaginal smear is a method to determine the estrus phase experienced by
mammals. This method is based on the stage of estrus characterized by vaginal
epithelial cells that undergo cornification due to increased estrogen levels (Hafez,
1968). Vaginal smears using the vaginal area as an area identification. Vaginal
mucosa taken for material identification, where epithelial cells and leukocytes
present in the vaginal mucosa. Identification forms of epithelial cells and leukocytes
can be demonstrated in a phase of the estrous cycle (Gilbert, 1994).
Estrous cycle is the period between the estrus phase next estrus. This cycle
only occurs in non-primate mammals. In primate mammals such as estrous cycle
called the menstrual cycle. In one there are four phases of the estrous cycle yairu
proestrus, estrus, metestrus, and diestrus. Each phase is associated with changes in
activity and changes in the structure of the ovary, uterus, and vagina. Two different
types of cycles are found in female mammals. Humans and many other primates
have a menstrual cycle (menstrual cycle), while other mammals have estrous cycle
(estrous cycle). Both cases occurred after the endometrium starts to thicken and
teraliri a lot of blood, because the endometrium to prepare the uterus to allow the
implantation of the embryo. The difference between this cycle involves the fate of
both the uterine lining if pregnancy does not occur. In the menstrual cycle, the
endometrium is shed from the uterus through the cervix and vagina known as
menstruation (Campbell, 2004).
GnRH binds GnRH receptors on cells to stimulate gonadotropic FSH and LH
release into the circulation. The pattern of secretion of GnRH induces the release of
cyclic LH on the lower level of FSH. FSH induces follicular growth and secretion of
estradiol and inhibin by the granulosa cells. After ovulation luteinised granulosa and
theca cells begin to produce high levels of progesterone, gonadotropin releasing
hormone associated with gonadal atrophy in mammals after GnRH immunization.
GnRH-BSA conjugate modulates the hormonal levels in female mice. GnRH-BSA
immunization cause irregular estrous cycles and lower concentrations of estradiol
and progesterone. GnRH-BSA conjugates have deleterious effects on reproductive
hormones and estrous cycle in female rats (Javid et al, 2010).
Ovulation in mammals is associated with lust for absorption of large amounts
of estrogen into the bloodstream occurs just before ovulation. Ovulator spontaneous
are animals that do not require the insertion of an ovulation, such as cows, sheep,
pigs, goats, dogs, primates, rats and guinea pigs. Induced ovulatory are animals that
require the insertion of a normal ovulation, such as cats, rabbits, ferrite, mink and
members of the camel family (Frandson, 1992).
Ovulation in female mammals usually occurs around the estrus phase.
Defined as the period of estrus female mammals accept male mammals to marry. The
interval between estrous phase to the next phase of the estrous cycle known as estrus
(Hafez, 1968). Female reproductive organ in mammals is the ovary. The ovary is
composed of parts of the stroma and the cortex. Part ovarian stroma contains
connective tissue. Part of ovarian cortex consists of epithelial cells making up a nest
egg. Primordial germ cells in the cortex contained oogonia. One follicle has 16
oogonia primordial germ cells. One cell will become oocytes and 15 other cells
become nurse cells. Nurse cells synthesize RNA and protein functions to the oocyte.
In mammalian oocytes covered by follicular cells (granulosa cells). Oocyte growth
causes granulosa cell count increased and formed a layered wrap called theca. Theca
layer is divided into two: the theca interna and theca externa. Theca interna produce
the hormone estrogen (Rugh, 1962).
 Oviduct, uterus, vagina, and accessory glands are also an important
reproductive organs of animals breastfeeding. Oviduct is an outgrowth of the
mullerian ducts, fallopian tubes starting osteum form of a funnel tube (where the
fertilization of an egg by sperm), and fallopian with its winding shape. The uterus in
mammals including gymnoarian types, ie almost all parts produces mucus to
facilitate passing through the oviduct to the egg reaches the uterus. The uterus is the
organ that serves for implantation and development of the embryo into a fetus or
generate wrappers (Orphans, 1982).
According to Rugh (1962), ovulation occurs in the estrous cycle. Estrus cycle
period is estrus, metestrus, diestrus and proestrus. Estrus period occurs in cycles and
in a series of sequential except for the period of anestrus season like sheep cattle and
horses. General characteristics estrus period are as follows:
1. Estrus occurs in 12 hours which is characterized by behavioral estrus
2. Metestrus occurred at 21 hours indicated ovulation and corpus luteum formation
3. diestrus occurs in 57 hours are marked with the proper functioning of the corpus
luteum
4. proestrus occurs in 3-12 hours marked follicular development is rapid.
Phase is being experienced in Rodentia may be determined by observation of
vaginal smear. The proportion of epithelial cells showed nucleus piknotik growing
rapidly until ovulation, while the presence of leukocytes no. Piknotik cells emerge
and evolve during ovulation finish that looks like a clot and folded into the cell, in
addition to the effects of the hormone progesterone characteristics began to be seen
(Winton & Bayliss, 1962).
 IV. CONCLUSION AND RECOMMENDATION

A. Conclusion

Based on previous results and discussion can be concluded as

follows:
1. Vaginal smears are used to identify the types of cells in the
vaginal smear, and to determine the phase - phase of the estrous
cycle that occurs the test animals. Cell types identified in vaginal
smears are epithelial cells and leukocytes.
2. There are four stages of the estrous cycle is proestrus, estrus,
and diestrus metestrus.
3. Cell types in the estrus phase of the estrous cycle in female
mammals is their ephitel terkornifikasi.
4. Practicum vaginal smear obtained for this time of estrus in
female mice phase characterized by epithelial terkornifikasi and
going for 18 hours.

B. Recommendation

1. As good as it is currently supplied practicum vaginal smears were already finished

so as to help groups that have managed to make the vaginal smear .
2. As good as it's aat practice um supplied four vaginal smears which consists of
phase proestrus, estrus, mesestrus / postestrus, and diestrus.
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musculus. Iranian Journal of Reproductive Medicine. 8 (2): 70-75.
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Frandson, RD 1992. Anatomy and Phisiology of Farm Animal. Lea Febigur,
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Taylor, Pamela. 1994. Practical Teratology. WB Saunders Co. London
Winton and Bayliss. 1962. Human Physiology. Brown and Company. Boston