Separation of sensitized and non-sensitized RBCs: sephadex-based cell-affinity adsorbents

In transfusion medicine, antibodies that cause RBCs positive DATs, may interfere with patients' phenotyping. Traditionally, these antibodies were removed using various antibody elution methodologies. However, the elution agents and conditions used have been only partially successful; and no one method is superior. The purpose of this study was to develop a general and efficient method to separate non-sensitized from sensitized RBCs using Sephadex-based cell-affinity adsorbents.

First, we coupled Sephadex support with Staphylococcal Protein G (SpG) with or without NHS. Then we simulated clinical conditions by mixing different ratios of sensitized and non-sensitized RBCs in vitro. Sensitized cells were prepared by mixing antibody with corresponding antigen-positive RBCs. Finally, we checked the sensitization status of absorbed RBCs after absorption with modified Sephadex support.

The number of sensitized RBCs bound to Sephadex-based cell-affinity adsorbents is approximately 5×10(8) RBCs/mL support. Activated Sephadex could separate sensitized from non-sensitized RBCs. Conclusion Sephadex-based cell-affinity adsorbents with an NHS spacer arm have bigger capacity for binding RBCs than unmodified Sephadex. The Sephadex-based cell-affinity adsorbents readily separate non-sensitized RBCs from sensitized RBCs, thus providing a new strategy to type the blood for transfused patients.