What is the Quadruple or Combined Test?

The quadruple or combined test is a screening test based on the analysis of several markers or chemicals in the mother’s blood. It is performed in the second trimester of pregnancy, between the 15th and 22nd weeks. The results are most accurate when done between 16 and 18 weeks.

The test results are combined with other factors such as the mother’s age, racial background, weight, diabetes status and gestational age, to arrive at a risk estimate for certain congenital defects.

Value of the Quadruple Test

It helps to estimate the risk of the following conditions in the fetus:

• Down syndrome or trisomy 21
• Open neural tube defects including myelomeningocele and spina bifida
• Trisomy 18

Implications

When the test is positive, it means the threshold value has been exceeded. This does not mean the baby has a birth defect, but that more testing is required to rule it out. The next step is normally an ultrasound scan. The first reason for this is to check the fetal age, since wrong dating of the pregnancy is the most common reason for a positive result. Secondly, it will show up many problems. If any of these are found, further testing and/or counseling may be required.

Individual Components

Alpha-Fetoprotein (AFP)

AFP is a glycoprotein that makes up the major part of the blood protein in the fetus. It is produced by the yolk sac at first and then by the liver. Its concentration is highest by 10-13 weeks and then slowly reaches a steady state at about 250 ng/mL. After 32 weeks of pregnancy it begins to fall again. Some amount of AFP leaks into the maternal blood and this is tested.

High Alpha-fetoprotein (AFP) levels are found in:

• Wrong dating of the pregnancy, when the fetus is older than is thought by the menstrual dates
• Multiple pregnancy
• Open neural tube defects as the AFP in the fetus passes through the defect into the amniotic fluid and the placental circulation, and from there to the mother’s blood
• Gut defects involving the abdominal wall, such as omphalocele and gastroschisis
• Esophageal atresia
• Congenital kidney defects
• Threatened abortion
• Fetal death

Low Alpha-fetoprotein (AFP) levels may occur in trisomy 21 and trisomy 18.

Estriol

This is tested as unconjugated estriol (uE3), which is the main form of the female hormone in pregnancy. It is produced in the placenta and it increases over the term of pregnancy.

Low levels of uE3 are found in:

• The trisomies
• When the fetus is thought to be older than it really is (wrong dates)
• Miscarriage
• Some genetic conditions such as X-linked ichthyosis

Human Chorionic Gonadotropin (hCG)

This is a glycoprotein with an alpha subunit which is shared by many other hormones, including LH, FSH and TSH. The beta subunit is most similar to that of LH and it gives hCG its unique function of maintaining pregnancy by helping the corpus luteum survive as a source of progesterone, until the placenta is established. The placenta then takes over the job of producing this hormone until it is time to deliver the baby. Thus the concentration of hCG is highest in the first trimester and doubles every 48 hours or so, beginning to fall at the end of the first 13 weeks.

High hCG levels occur in Down syndrome and in multiple pregnancy, but low levels in trisomy 18. It may also be falsely high if the fetus is thought to be older than it really is, due to wrong dating, because this test is normally performed in the second trimester when the levels normally start to fall. Triploidy, hydrops fetalis, and fetal loss, are other reasons for a high hCG level.

Inhibin A

This glycoprotein comes from the ovarian granulosa cells and the testicular Sertoli cells, which are nutritive in function. It has an alpha and a beta subunit. The latter exists in different of the inhibin hormone. Inhibin is important in regulating ovarian hormone production by feedback acting upon the pituitary and hypothalamus, as well as determining various aspects of fetal development. It is produced by the placenta in increasing concentrations as pregnancy progresses and as the fetus grows larger.

Abnormalities in inhibin A levels mirror those of hCG and have the same significance, but help to make the diagnostic value of the quadruple test more specific and sensitive.