“The cell-free fetal DNA test is a type of prenatal genetic test used for screening various genetic conditions associated with a fetus.” 


The cell-free DNA is different from nuclear DNA,

“The cell-free DNA are the fragments of nucleic acid floating in maternal blood used for prenatal genetic screening. “

In the present article, we will discuss about the futuristic prenatal genetic testing technology- the cell-free fetal DNA test.

What is cell-free DNA? 

The cell-free DNA is freely floating smaller fragments of fetus DNA present in the maternal blood. After the 10th week of gestation, the shed off of trophoblastic cells in maternal blood released foetal DNA fragments.

In the year 1997, Lo YM, Tsui NB, Chiu RW and co-workers developed a cell-free DNA testing method. It’s also known as noninvasive prenatal genetic testing or cell-free fetal DNA testing, abbreviated as cfDNA or cffDNA.

Several chromosomal and gene mutations can identify using the present method. Further, the sex and the Rh factor of the fetus can also determine. 

How it’s done? 

The procedure of it is simple, yet, complicated. Approximately 10 ml of blood is drawn from the mother (Yes, you heard it right!). The serum is isolated using the high-speed centrifugation, keep in mind not to speed high enough to damage the DNA. 

The cell-free DNA fragments are smaller than nuclear DNA and other cell organelles. Therefore it floats in serum. The isolated serum is proceeding for DNA extraction. 

Here, maternal DNA doesn’t require, therefore, the pallets are of no use. Now using the cell-free DNA isolation kit or a standardized protocol, if you have! DNA isolation is performed

Notably, if your protocol is standardized, a small amount of formaldehyde can increase the yield of the DNA. Avoid if you are using the kit. 

Once the DNA is extracted, it is stored at 4°C and processed for polymerase chain reaction or DNA sequencing. 

The process of the cell-free DNA test.

Why it is needed?

Cell-free fetal DNA testing fulfils two main objectives for pregnancy screening. It’s an effective non-invasive option and used for genetic screening. 

The invasive method for sample collection has several limitations,

  • Obviously it is painful- sample is collected by holing or cutting tissue. 
  • The site of sample collection may be infected with some microorganisms. 
  • Or some time extreme pain, swelling or uncontrolled bleeding (in case of haemophilia) happens. 

Therefore, invasive techniques such as amniocentesis and chorionic villi sampling are not recommended always. Plus, both the present techniques are riskier. miscarriage chances are always high in it.

In the cell-free fetal DNA testing, we are not touching the fetus and hence it is safer. 

It is applicable to a wide range of genetic screening. It’s used in the screening of common trisomies- 13, 18 and 21(Down syndrome). Furthermore, It is also applicable for single-gene disorders like sickle cell disease, thalassemia or cystic fibrosis

Besides this, other chromosomal aberrations like aneuploidies and major deletion, sex of the fetus and Rh factor can be determined. However, it isn’t working finely for complex genetic disorders.

read more on trisomies:

  1. Trisomy 18
  2. Trisomy 21 (Down Syndrome)
  3. Trisomy 13(Patau Syndrome)

When to go for it? 

Any pregnant women can go for cell-free DNA testing. Doctor’s advice is mandatory before testing it. 

Notably, in the below-enlisted conditions women can go for it: 

  • Previously genetically abnormal pregnancy 
  • Stillbirth 
  • Carrier father or mother 
  • Family history of genetic disorders 
  • Two or more miscarriage 
  • Present fetus screened with any physical abnormalities. 

Nonetheless, the doctor’s advice and genetic councillor’s recommendation is required.

When not to go for it? 

Not all cases are suitable for cfDNA testing. It is less effective in the case of females with twin pregnancy, obesity, in vitro fertilization, fertilized with a donor, taking certain medications for blood thinner, diabetes or pregnancy less than 10 weeks.

Generally, the doctor avoids cfDNA testing in these cases. Still, if the female has any indications of genetic abnormality, it can be done. 

What are the risks?

No physical risk is associated with female or her fetus. So no need to worry about it, nonetheless, cell-free fetal DNA testing has several limitations. In some of the cases, it might cause anxiety in females. 

What are the limitations? 

Cell-free DNA fragments are just fragments of DNA, not the entire genome of the fetus. Thus all type of genetic abnormalities (chromosomal as well as DNA) can’t be screened using the present technique. 

Moreover, it is not considered as a diagnostic method, unlike genetic testing using amniocentesis. 

It is more a kind of screening method, the final results of the cfDNA test must be validated using amniocentesis or chorionic villi sampling.

The negative results don’t mean that the fetus is absolutely fine. Whereas the positive results even don’t mean that the fetus is abnormal. 

As we said above, genetic testing using any of the sample mentioned above must perform before confirming results. 

In addition to this, one major limitation of the cell-free fetal DNA testing is the fragments of DNA. One who has done practical work on it can only know about this problem. 

What DNA fragments we get, are not always what we want. What does it mean?

For instance, if you want to screen the beta-globin gene for sickle cell disease, all the time you can not obtain the beta-globin gene fragments. I mean, cfDNA are just random fragments of DNA, not an entire genome. 

Hence it is possible that you want to screen beta-thalassemia and you don’t get a fragment of it. 

What do the results indicate? 

The results of the cfDNA testing are similar to genetic testing. The patient can expect results in 10 days. The major concern of the results taken into account when it’s positive. 

If screened positively, immediately a patient has to go for the prenatal genetic testing. 

For example, the result indicates down syndrome, immediately, AF or CV sample is collected and send for cell culture in order to perform chromosome test

Besides all these, one of the major advantages of the present test is in the determination of sex. The sex of the fetus can be determined using cfDNA testing.  

Note: prenatal sex-determination is an offendable act in countries like India. 

Prenatal genetic testing helps in identifying genetic conditions associated with the fetus. Generally, these conditions are congenital and non-curable.

Also, it burdens families emotionally, economically and socially. 

Hence it is advisable to abort an abnormal child if parents willing to do. It helps in making a decision related to their fetus. 

In addition to this, carriers and normal are identified and can live a healthy life. Therefore in any of the condition, cell-free DNA testing is beneficial for the patient. 

Still, the present testing method isn’t fully ready to use in the diagnosis, yet. It’s even too expensive. A middle-class family can’t afford it. 

I had worked in the prenatal genetic testing department in past. I had also performed several experiments for standardizing cell-free DNA extraction protocol. Trust me it is too tough to isolate DNA from it. 

Only a couple of times, I had got some amount of DNA but not the fragment I want to study. Still, I had tried to sequence those fragments which might be some DNA sequences of X-chromosome.

The results were not conclusive. 

So, the biggest problem related to the present method is isolating DNA. I am requesting all the scientists, if you have some protocol related to cfDNA, please share it with others.

It will help to establish this revolutionary technique for effective prenatal diagnosis. 

Read more:

What is DNA sequencing?

Conclusion: 

Less published data on cell-free DNA test is available. Further, no standardised protocol for DNA extraction for cfDNA is available too, and even if it is, scientists are trying to sell it on a high price to DNA extraction kit making companies.  

Notwithstanding, because of it’s outstanding applications, it can be a revolutionary technique in future.