Optimizing Kappa-Deleting Recombination Excision Circles (KREC) Cut-Off Values in Russian Newborn Screening Program:...

Newborn screening (NBS) for primary immunodeficiencies (PIDs) increasingly incorporates kappa-deleting recombination excision circles (KREC) to enable early detection of B-cell lymphopenia, particularly agammaglobulinemia (AGG). However, optimal KREC cut-off values remain poorly defined, limiting sensitivity and contributing to high false-positive rates. We evaluated KREC-based NBS using data from a nationwide Russian program that screened more than 3.4 million newborns. KREC measurements from 66 patients with genetically confirmed AGG and 534 newborns with initially abnormal screening results but normal immunophenotyping were analyzed using statistical and ROC-based approaches. By using a KREC cut-off of 100 copies per 10⁵ cells in the first NBS sample and a KREC cut-off of 250 copies per 10⁵ cells after a polymerase chain reaction (PCR) test on a second dried blood spot (DBS) collected approximately 19 days later, the detection of classical and atypical X-linked AGG, other forms of AGG, and additional PIDs associated with B-cell lymphopenia was significantly improved. Retrospective analysis of a pilot cohort (n = 202,908) indicated that, if a KREC cut-off of 250 copies per 10⁵ cells was applied with the second PCR test, this adjustment would increase the proportion of screen-positive newborns to 2.75‱. To reduce the resulting referral burden, we assessed third KREC testing of a third DBS sample approximately 2 months after birth and showed that persistent KREC reduction distinguishes AGG from transient B-cell lymphopenia. We propose a revised three-step NBS algorithm incorporating a third PCR test with a cut-off of 850 copies per 10⁵ cells. This strategy reduces unnecessary referrals while maintaining diagnostic sensitivity and clinical safety, supporting implementation of adaptive, multistep NBS approaches.