Diagnosis of PKU

Newborns who are diagnosed with phenylalanine hydroxylase deficiency may show no initial symptoms or signs.

Over time they may develop symptoms of phenylalanine build-up in the body.

In the body phenylalanine is metabolized by hydroxylation of phenylalanine to tyrosine by phenylalanine hydroxylase (PAH).

When there is phenylalanine hydroxylase deficiency the blood and plasma levels of phenylalanine rise. This is more often than not diagnostic of the condition of PKU or phenylketonuria.

Plasma levels of phenylalanine

Plasma levels of phenylalanine are consistently over 120 µmol/L (2 mg/dL) before therapy is initiated.

A Guthrie card bloodspot is used. The newborn’s heel is pricked with a needle to obtain a spot of blood over the specially treated diagnostic card.

If the levels of phenylalanine is high in blood it shows up in the card. In case of a first positive test, a second test is needed.

Once the test shows positive second time, further tests are needed to confirm the diagnosis. (1, 2, 3, 4)

Tests at birth for PKU

Tests at birth include (1) –

  • Guthrie card bacterial inhibition assay (BIA) – this is an inexpensive, simple and easy-to-use test that is applied routinely.
  • Fluorometric analysis – This requires machinery that tests the blood for raised levels of phenylalanine. This shows more accurate results than the BIA.
  • Tandem mass spectrometry (MS) – this test has similar reliability as fluorometric analysis. It can also find the levels of tyrosine.

    Phenylalanine is ultimately broken down into tyrosine with the help of phenylalanine hydroxylase. In PKU there is a deficiency of phenylalanine hydroxylase. Thus this test gives a more accurate picture of the condition.

Diagnosis of carriers of PKU

Sometimes carrier status of the disease among parents and in a baby may also be detected using biochemical parameters rather than genetic testing.

This is done using ratios of Phenylalanine and tyrosine in blood.

There is a normal variation of this ratio in blood based on the time of the day called the circadian rhythm.

Thus, this test needs to be performed before noon after a normal breakfast. The results may also vary at various phases of a woman’s menstrual cycle and may not be accurate during pregnancy.

Molecular testing for PKU

Molecular testing includes looking at the phenylalanine hydroxylase deficiency via the activity and functioning of the PAH gene on the 12th chromosome.

There are between 1 and 15 faults or mutations. These can predict nearly 30-50% of genetic defects in the PAH gene. A target identification looks at these faults specifically.

There may also be scanning for the mutation that looks at all defects at the location, or sequencing of the gene for mutation detection.

Sometimes genetic lookouts may be for deletions or duplication of segments of DNA at the gene site as well. These can detect accurately the mutation and the risk carried by an individual.

Some parents may need genetic diagnosis of carrier status for PKU before conception especially if the condition has been seen in the family. (1)


  1. http://www.ncbi.nlm.nih.gov/books/NBK1504/
  2. https://learn.genetics.utah.edu/
  3. http://www.genome.gov/25020037
  4. https://www.ncbi.nlm.nih.gov/

Further Reading

Last Updated: Jun 12, 2023

Dr. Ananya Mandal

Written by

Dr. Ananya Mandal

Dr. Ananya Mandal is a doctor by profession, lecturer by vocation and a medical writer by passion. She specialized in Clinical Pharmacology after her bachelor's (MBBS). For her, health communication is not just writing complicated reviews for professionals but making medical knowledge understandable and available to the general public as well.


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