Preparation of clinical-grade WBCs using leukocyte reduction filters.

OBJECTIVE

Our goal was to develop a simpler and less expensive method of obtaining human clinical-grade WBCs using an alternative method to continuous leukapheresis. Our purpose for the WBCs is to arm them with rabbit anticancer antibodies for a phase I clinical trial.

METHODS

Using leukocyte reduction filters (LRFs) discarded from the blood bank, we evaluated multiple variables to maximize recovery of WBCs with the lowest contamination of RBCs. Using an optimized protocol, full-scale runs according to FDA current Good Manufacturing Practice (cGMP) standards were completed with immediate filtration of blood obtained from donors participating in our study.

RESULTS

Forward flushing of the filter removed 85% to 95% of residual RBCs and platelets. When backward flushed with 800 mL, 95% of the WBCs recovered were contained in the first 400 mL. The number of recovered WBCs was in the range of 166-211 million/100 mL filtered blood. Subpopulations of WBCs recovered from the LRFs were in the same proportion as the donors' whole blood. Viability of recovered WBCs was 96-99%. Exogenous rabbit antibodies bound well to the recovered WBCs and were retained for at least 5 h without significant reduction. Three full scale runs of WBCs recovered from donor blood filtered through the LRF met all FDA specification of sterility, endotoxin levels, viability and stability.

CONCLUSION

Using LRFs, high quality clinical grade WBCs are readily obtained in quantities of 0.2 to 1.2 billion cells from 100 mL to 450 mL (1 unit) of whole blood.