*3.3.1. Invasive methods to obtain embryo-fetal cells*

The methods for obtaining embryonic or fetal cells are chorionic villus sampling, amniocentesis and cordocentesis.

**Chorionic villus sampling** (CVS) is a method that allows prenatal diagnosis in the first trimester of pregnancy, typically at 10–12 weeks of gestation. It is done transabdominal or transcervical, under ultrasound guidance. The samples obtained contain trophoblast cells (derived from fetus) and maternal decidua cells. The last must be removed before analysis. The major advantages of this technique are reduced maternal risk generated by termination of pregnancy and a limited emotional trauma on the patient. The risks of this method are injury of the embryo (especially limb abnormalities in early procedures), miscarriage, bleeding or embryonic infection [102, 103]. The risk of miscarriage after performing chorionic villus sampling was initially estimated at 2–3%. Recent data disproves this risk, showing that the real risk is about 0.22% [104].

**Amniocentesis** is performed from the 16th week of gestation (rarely sooner) and involves the transabdominal extraction of 10–20 ml of amniotic fluid, under ultrasound guidance. The method has a low risk of miscarriage and other incidents. The risk of miscarriage was initially estimated at 1%, but recent data disproves this risk, showing that the real risk is about 0.11%. There is also a risk of amniotic fluid leakage and some rare complications (placental hemorrhage, intra-amniotic infection, abdominal wall hematoma and fetal lesion). A major disadvantage of this method is that it provides the final results of prenatal diagnosis in the second half of the pregnancy [103–105].

**Cordocentesis** is a method that consists of transabdominal or transvaginal puncture of the umbilical cord under ultrasound guidance in order to obtain small amounts of fetal blood. The method is performed after week 20 of pregnancy and has a reduced applicability in the diagnosis of chromosomal diseases. It is usually used to diagnose fetal blood diseases. Risk of fetal loss is high, about 3%. The major advantage of the diagnosis of chromosomal disorders is associated with the best results of blood cells culture in comparison with amniotic or chorionic cells [106].

repeated after amniocentesis or cordocentesis to determine real fetal chromosomal formula. Another problem associated with CVS is the failure of culture that requires the repeat of CVS

Prenatal Biochemical and Ultrasound Markers in Chromosomal Anomalies

http://dx.doi.org/10.5772/intechopen.73604

383

Fetal karyotype analysis, using cells obtained by amniocentesis, requires a step of 10–14-day cell culture. Culture technique and processing steps are similar to those used in the case of trophoblastic cells. Amniotic cultures can be done in situ (allow differentiation between mosaics and pseudomosaics) or in culture flasks. In the last case, at the end of the cultures, the separation of cells from the vessel is obtained using an enzymatic treatment. The contamination with maternal cells is insignificant and the quality of prepared chromosomes is better in

Molecular techniques for prenatal diagnosis of chromosomal disorders were introduced in the medical practice in the last 25 years with the aim of improving the resolution of chromosomal detection and to eliminate the major inconvenient of classic cytogenetic techniques the requirement of cell culture. Such techniques are fluorescence in situ hybridization (FISH), quantitative fluorescent polymerase chain reaction (QF-PCR), multiplex ligation probe ampli-

FISH technique allows a hybridization between a fluorescent probe and a complementary DNA sequence present on the target chromosome. In prenatal diagnosis this method has been developed in order to identify the aneuploidy of chromosomes 21, 18, 13, X and Y (the most common chromosomal abnormalities) (**Figure 7**). The probes for chromosomes 13 and 21 are locus specific while the rest are centromeric. The method is done on uncultivated interphasic amniocytes. The final results are obtained in 24–48 hours after amniocentesis which allows maternal anxiety relief [108–110]. The results are obtained after the evaluation of minimum 50 cells. The main advantages are faster results, high specificity and sensibility (close to 100%) and the absence of cell culture. The inconvenients are impossibility of detection of blood contamination, bad quantification of chromosomal mosaics and impossibility of detection of structural abnormalities of chromosomes investigated. FISH can be done also for the evaluation and elucidation of uncommon structural abnormalities: microdeletion syndromes, cryptic or subtle duplications and translocations, complex rearrangements and marker chro-

QF-PCR method allows the detection of major prenatal numerical chromosome disorders within 24–48 hours. The method identifies polymorphic chromosomal specific repeat sequences (short tandem repeats—STRs) that are amplified by the PCR using fluorescent

fication (MLPA) and array-comparative genome hybridization (CHG).

or the application of amniocentesis in the second trimester of gestation [103, 107].

*3.3.2.1.2. Second-trimester cytogenetic analyses*

comparison with chorionic villus cells [103, 107].

*3.3.2.2. Molecular techniques*

*3.3.2.2.1. Fish*

mosomes [111].

*3.3.2.2.2. QF-PCR*
