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The prediction that AFC culture would largely be replaced by CVS (see Chapter 9) analysis has not come true. Coelocentesis at 6–10 gestational weeks was proposed as an alternative to both CVS and amniocentesis but did not achieve wide acceptance.⁵²⁰ The use of early amniocentesis (9–14 gestational weeks) has been limited by increased fetal risk (see Chapter 9), failure to obtain fluid and operator inexperience rather than by failure to grow cells from such early specimens.^{521, 522} Although never widely adopted, rapid and quantitative PCR methods have been employed to identify the common autosomal and sex chromosome aneuploid states.^{523, 524} Interphase fluorescent in situ hybridization (FISH) technology was also predicted to replace cell culture and metaphase karyotype analysis. This prediction has not been realized except for occasional cases of malformations identified by ultrasound examination and then confirmed as aneuploid using FISH methods.^{525, 526} Most of these cases are still confirmed by cell culture and karyotype analysis, albeit after pregnancy termination. Interphase FISH analysis can identify 90–95 percent of the clinically relevant autosomal and sex chromosomal aneuploidy, although up to 25–30 percent of all cytogenetic changes (e.g. mosaics and balanced translocations) are detected only by G‐banded metaphase analysis.^527–534 Prenatal chromosomal microarray is gaining rapid acceptance as an alternative to conventional chromosome analysis, and is now recommended as the first‐tier test when ultrasound examination identifies malformations.^535–537

In cases of bladder outlet obstruction, it is possible to obtain fluid and cells for culture from the fetal bladder. In one study, karyotype analysis was successful in 95 percent of 75 samples, with six chromosome abnormalities identified.⁵³⁸ AF may not be accessible in cases of severe cystic hygroma, pleural effusion, renal agenesis, or bladder outlet obstruction. Cases presenting with severe cystic hygroma, ascites, or pleural effusion are at significant risk of having a chromosome abnormality, especially trisomy 21 or monosomy X.⁵³⁹ Fluid drawn from any of these sources will contain cells that can be cultured like amniocytes and usually also include lymphocytes that can be cultured with phytohemagglutinin (PHA).^540–542

The increasing acceptance and sophistication of maternal serum screening tests and high‐resolution prenatal ultrasonography identified a greater number of at‐risk pregnancies, especially in women under age 35. Before the year 2000, this resulted in an increase in the number of amniocentesis procedures. However, since the early 2000s, the number of amniocenteses for karyotype studies has been falling. At first this was because women over age 35 used screening‐based risk figures to decide against prenatal diagnosis if their risks were deemed to be reduced based on triple, quadruple, or more complex screening tests that combine information from the first‐ and second‐trimester serum assays and ultrasound examinations (see Chapter 6). With each additional chemical component of maternal serum screening came a higher rate of detection and a lower false‐positive rate – hence fewer women were “screen positive” and many elected to forego amniocentesis.^{543, 544} As fetal DNA isolated and sequenced from maternal serum gains increased acceptance,⁵⁴⁵ the number of prenatal karyotype studies will continue to decline. False‐positive and false‐negative results will likely keep these methods as screening rather than diagnostic tests.^{546, 547}