Pregnancy Tests

Pregnancy test is the test used to detect or confirm pregnancy. The basis of pregnancy tests is to determine the presence of the human chorionic gonadotropin (hCG) in the urine of woman suspected for pregnancy. Both biological and immunological tests are available to determine the presence of hCG in the urine of the pregnant woman.
Pregnancy Tests


These tests are performed by using experimental animals. The biological tests for pregnancy can be performed only after 2 or 3 weeks of conception so that, the concentration of hCG in urine is sufficient to show the result.


Aschheim-Zondek test was the first test invented for confirming the pregnancy. It depends upon the ovarian changes in immature mice caused by hCG. The immature mice do not ovulate naturally. Ovulation occurs only if hCG is injected. 2 mL of urine from the woman suspected for pregnancy is injected daily for 2 days into the immature mice. 5 days after injection of urine, the mice are killed. The ovaries are examined for the presence of corpora lutea (plural for corpus luteum) and hemorrhages, which indicates ovulation. Ovulation is due to the presence of hCG in urine.


Kupperaman test is the modification of Aschheim- Zondek test, in order to save time. In this, an immature

rat is used instead of immature mice. About 2 mL of urine is injected subcutaneously into immature rat and ovarian changes are observed after 6 hours. If the urine is injected intraperitoneally, the ovarian changes can be observed within 2 hours.


In this test, 10 to 15 mL of urine is injected intravenously into rabbit and ovulation is observed by examining the ovaries after 48 hours.


In this test, about 20 to 30 ml of urine is concentrated and injected into the dorsal lymph sac of South African toad, Xenopus levis. If hCG is present in the urine, it causes ovulation after 12 hours.


In this test, 2 mL of urine is injected into the male amphibian (toad or frog). hCG in urine causes expulsion

of spermatozoa within 2 hours. Biological tests are outdated after the development of immunological tests.

Disadvantages of Biological Tests

Biological tests for pregnancy are replaced by immunological tests because of several disadvantages:

1. The biological test require animals

2. Tests can be performed only after 2 to 3 weeks of pregnancy so that sufficient quantity of hCG is excreted in urine

3. Results are not obtained quickly; one has to wait for 2 to 48 hours

4. Tests involve tedious procedures such as sacrificing the animals.


Presence of hCG is also determined by using immunological techniques. Immunological tests

are based on double antigen-antibody reactions. Commonly performed immunological test is known as

Gravindex test.


Principle is to determine the agglutination of sheep RBCs coated with hCG. Latex particles could also be

used instead of sheep RBCs.


1. Antiserum from Rabbit

Urine from a pregnant woman is collected and hCG is isolated. This hCG is injected into a rabbit.

The rabbit develops antibodies against hCG. The antibodies are called hCG antibody or anti-hCG. The

rabbit’s blood is obtained and serum is separated. The serum containing hCG antibody is called rabbit

antiserum or hCG antiserum. It is readily available in the market.

2. Red Blood Cells from Sheep

RBCs are obtained from sheep blood and are coated with pure hCG obtained from urine of the pregnant women. Nowadays, instead of sheep RBCs, the rubberized synthetic particles called the latex particles are used.

3. Urine

Fresh urine sample of the woman, who needs to confirm pregnancy is used for Gravindex test.


1. One drop of hCG antiserum is taken on a glass slide. One drop of urine from the woman who wants

to confirm pregnancy is added to this and both are mixed well.

2. Now, one drop of latex particles is added to this and mixed.


Result is determined by observing the agglutination of latex particles added to mixer of hCG antiserum and woman’s urine.

Absence of Agglutination of Latex Particles

If hCG is present in urine, it is agglutinated by antibodies of antiserum and all the antibodies are fully used up.

 Agglutination of hCG molecules by antibodies is not visible because it is colorless. Later when latex particles are added, these particles are not agglutinated because, free antibody is not available. Thus, absence of agglutination of latex particles indicates that the woman is pregnant.

Presence of Agglutination of Latex Particles

If urine without hCG is mixed with antiserum, the antibodies are freely available. When latex particles

are added, the antibodies cause agglutination of these latex particles. Agglutination of latex particles can be seen clearly even with naked eye. Thus, presence of agglutination of latex particles indicates that, the woman is not pregnant.



1. Immunological tests are more accurate

2. Result is obtained quickly within few minutes

3. These tests can be carried out very easily. The procedure is not cumbersome, as in the case of biological tests

4. Immunological tests can be performed on 5th day of conception. By biological methods, the tests can

be performed only after 2 or 3 weeks of conception. It is because, the concentration of hCG required

for producing changes in the animals is excreted in urine only after 2 or 3 weeks of pregnancy

5. Recently available immunological tests are more sensitive and involve single step method. Test kit

is available in the form of cards. These pregnancy test cards can be used even in the first few days

of conception. Most sensitive test can detect hCG level as low as 20 mIU/mL.


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