# Integrating RHD Genotyping for More Accurate Rh(D) Antigen Phenotyping: A Retrospective Study

## TL;DR

This study shows that using RHD genotyping improves the accuracy of Rh(D) blood type classification, reducing risks and optimizing blood supply in transfusion medicine.

## Contribution

The study demonstrates the clinical impact of integrating RHD genotyping for resolving Rh(D) discrepancies and optimizing blood transfusion practices.

## Key findings

- Weak Rh(D) variants were most common in pediatric patients, with Weak Type 2 being predominant.
- Partial Rh(D) phenotypes were more prevalent among Black individuals.
- Genotyping reclassified some Rh(D)-negative patients as Rh(D)-positive, reducing unnecessary Rh Immunoglobulin use.

## Abstract

Background and Objectives: The Rh blood group system is highly polymorphic, and accurate classification of Rh(D) variants is critical in transfusion medicine to prevent alloimmunization and optimize blood utilization. Despite the advances in conventional serologic testing, weak and partial Rh(D) phenotypes still remain challenges in Transfusion Medicine practice. The objective is to implement and assess the impact of RHD genotyping in classifying Rh(D) antigen status. Materials and Methods: We conducted a retrospective study at the University of South Alabama Medical Center and Children and Women’s Hospital between 1 January 2023 and 31 December 2024 to assess the impact of RHD genotyping in cases with discrepant Rh(D) typing, Rh(D)-positive patients with anti-Rh(D) antibodies, and neonates with positive weak Rh(D) tests. ABO and Rh(D) antigen typing was performed on 12,994 patients, including 3767 newly tested individuals. Weak Rh(D) testing was performed on newly tested individuals using automated microplate direct agglutination, followed by molecular genotyping. Results: Among the 25 patients with weak or discrepant Rh(D) phenotypes, weak Rh(D) variants were observed in 52% of cases, with Weak Type 2 being the most common, particularly in pediatric (age < 18 years old) patients. Partial Rh(D) phenotypes were identified in 40% of cases, predominantly among Black individuals. Three patients were reclassified as Rh(D)-positive based on genotyping and received 615 Rh(D)-positive RBC units without evidence of alloimmunization, while four patients were confirmed at risk of alloimmunization and remained classified as Rh(D)-negative. Fisher’s exact test demonstrated a significant association between ethnicity and Rh(D) classification (p < 0.01), and the McNemar exact test confirmed a significant reclassification of cases from Rh(D)-negative to Rh(D)-positive (p < 0.01). Conclusions: RHD genotyping enhances the accuracy of Rh(D) antigen classification, mitigating alloimmunization risks and the unnecessary use of Rh Immunoglobulin and optimizing blood product utilization. The reclassification of patients to Rh(D)-positive alleviates pressure on Rh(D)-negative blood supplies, particularly during critical shortages. These findings underscore the necessity of integrating molecular RHD testing into routine transfusion medicine practices to improve patient safety and resource management.