Screening for primary immunodeficiency diseases and follow-up testing

Oct. 24, 2016

Until recently, there had been no newborn screening test for primary (or genetically-determined) immunodeficiency diseases. That changed in 2010 when the United States Secretary’s Advisory Committee for Heritable Disorders of Newborns and Children (SACHDNC) voted unanimously to add screening for severe combined immunodeficiency (SCID) and other causes of T cell lymphopenia to the recommended uniform screening panel (RUSP) for newborns.1 The Secretary of the U.S. Department of Health and Human Services approved this recommendation soon after, and there are currently 39 states screening for these conditions.2 While this is a first for primary immunodeficiency, considering the available molecular technology, it is just a matter of time before screening tests for many other genetic defects in the immune system will be developed.

Screening for SCID

The screening test for SCID and other causes of T cell lymphopenia is performed on dried blood spots, just as all other RUSP tests are. However, since performance of this test requires molecular technology, many states were slow to adapt this assay because they lacked personnel with expertise to perform it. The procedure detects T cell receptor excision circles (TRECs), which are pieces of DNA that are excised and form circles within thymocytes when the T cell receptor genes undergo rearrangement. These cells subsequently leave the thymus and enter the peripheral blood. TRECs are quantified on neonatal dried blood spots by the real-time polymerase chain reaction (RT-PCR).3 Normal newborns have a very large number of TRECs, whereas SCID infants—as well as those with other defects resulting in low T cells numbers—have very low numbers or no TRECS.

The experience to date has shown that the TREC assay has detected a number of other conditions in addition to typical and leaky SCID and Omenn’s syndrome (an SCID variant).2 These include, among others, the DiGeorge anomaly, Charge syndrome, ataxia telangiectasia, trisomies 18 and 21, and many other genetic syndromes with impaired T cell development. In addition, there are other conditions with secondary T cell impairment, such as cardiac anomalies, multiple congenital anomalies, gastrointestinal anomalies, etc., that have been identified by finding abnormally low numbers of TRECs. Even more exciting is the fact that idiopathic T cell lymphopenic conditions have been discovered through this testing which undoubtedly represent new genetic defects heretofore unknown.

Determining the diagnosis

The above information emphasizes the importance of referring infants with positive screens to an immunologist to determine the correct diagnosis before treatment is even considered. The initial follow-up testing is performed by flow cytometry on anti-coagulated venous blood obtained when the infant is seen promptly in follow-up. A complete blood count and manual differential should also be performed. The cell surface markers important to test for in flow cytometry (at a minimum) are CD3, CD4 with CD45RA (a marker for naïve T cells) plus CD62L (or CD31), CD45RO, CD8, CD16 and CD20. The co-expression of CD45RA plus CD62L on CD4 positive helper T cells defines a recent thymic emigrant. SCIDs, leaky SCIDs, and babies with Omenn’s syndrome lack CD4 positive T cells that co-express these markers, whereas normal newborns have a very high number of recent thymic emigrants.

If there are CD3 positive T cells present in the screen-positive newborn, there is a possibility they could be maternal, because maternal T cells enter the fetal circulation during pregnancy and, if the fetus has SCID, he or she would not be able to reject the mother’s T cells. If that is the case, there could be CD4 positive T cells that co-express CD45RA but also other CD4 positive T cells that co-express CD45RO ( a marker for memory T cells), as is seen in normal adults. The presence of maternal T cells confirms the diagnosis of typical SCID.

Another confounding diagnostic problem is pre-maturity. Whether it is because the blood for the filter paper spots was collected in a neonatal intensive care unit where most infants would have central lines kept open by heparin locks (heparin interferes with PCRs) or just extreme prematurity, premature infants have a high incidence of false-positive TREC screens. It is important to recognize this and repeat the flow cytometry in a week or two. Finally, the most important follow-up testing is that of T cell proliferation in response to mitogens such as phytohemagglutinin (PHA). If there is no response to PHA, the infant is highly likely to have SCID.

Treatment considerations

The primary care physician who sees the screen-positive infant should be advised to keep him or her away from sick contacts, including those in the waiting room of a typical office. The infant should not attend daycare or similar activities where he or she will engage with other children, or, more generally, be taken to public places. Usually such infants are well at that point and do not need to be sent to an ER or hospital, where they could contract a nosocomial infection. In addition, they should not be given any live vaccines, such as rotavirus vaccine.

When the doctor sees the infant, inquiry should be made about a family history of immunodeficiency, and the infant should be examined to determine if there are physical features consistent with DiGeorge anomaly. If that is the case, testing for 22q11 should be initiated. The primary care physician should arrange to have the infant seen by an immunologist as soon as possible.

Treatment for SCID is a hematopoietic stem cell transplant4 (HSCT) or gene therapy. Treatment for the other conditions detected by TREC testing is neither of these—hence the importance of establishing the correct diagnosis before sending the infant for a transplant. If there is no transplant center in the state in which the infant is born that has experience in transplanting SCID infants, the SCID infant should be referred out of state to a center that does. Typical SCIDs do not need pre-transplant chemotherapy prior to HSCT because they are unable to reject a transplant. Pre-transplant chemotherapy can be highly toxic to newborns with SCID, and many transplant centers use these agents regardless of the condition they are treating. Thus, it is very important that these infants are referred to a center with considerable experience in transplanting SCID infants.


  1. Buckley RH. The long quest for neonatal screening for severe combined immunodeficiency.
    J Allergy Clin Immunol. 2012;129(3):597-604.
  2. Kwan A, Abraham RS, Currier R, et al. Newborn screening for severe combined immunodeficiency in 11 screening programs in the U.S. JAMA. 2014;312(7):729-738.
  3. Chan K, Puck JM. Development of population-based newborn screening for severe combined immunodeficiency. J Allergy Clin Immunol. 2005;115(2):391-398.
  4. Buckley RH. Transplantation of hematopoietic stem cells in human severe combined immunodeficiency: longterm outcomes. Immunol Res. 2011;49(1-3):25-43.

Rebecca H. Buckley, MD, is J. Buren Sidbury Professor of Pediatrics and Professor of Immunology, Duke University School of Medicine.