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Effects of Methylene Blue Stabilizer on In Vitro Viability and Chemotaxis of ^99mTc-Exametazime-Labeled Leukocytes

Nuclear Medicine, Department of Radiology, and Section of Biostatistics, Department of Health Sciences Research, Mayo Clinic, Rochester, Minnesota

This study was conducted to evaluate the effects of methylene blue/phosphate buffer stabilizer on the labeling efficiency, cell membrane integrity, chemotaxis, and in vitro stability of leukocytes labeled with ^99mTc-exametazime. Methods: Whole blood (480 mL) was obtained from each of 3 healthy donors and divided equally into eight 60-mL aliquots. Two aliquots from each of the 3 subjects were used as a control (i.e., leukocytes were not labeled with either nonstabilized or stabilized ^99mTc-exametazime) in this study. The remaining 6 aliquots from each of the 3 subjects were divided equally into the following formulation groups: (a) leukocytes labeled with nonstabilized ^99mTc-exametazime, (b) leukocytes labeled with stabilized ^99mTc-exametazime containing 250 µg methylene blue, and (c) leukocytes labeled with stabilized ^99mTc-exametazime containing 500 µg methylene blue. Duplicate samples were evaluated to confirm the repeatability of the study. Six samples were studied under each experimental condition (i.e., control, a, b, and c groups). The cell membrane integrity and chemotatic capacity of leukocytes were measured at 1 and 3 h after preparation, whereas a complete blood count was obtained at 3 h after preparation. The labeling efficiency was calculated immediately on conclusion of cell labeling, whereas the in vitro stability of the radiolabeled leukocytes was evaluated at 3 h after radiolabeling. Results: None of the samples showed trypan blue-stained cells. For the chemotactic index, no statistically significant differences were noted between the 3 labeling formulations at 1 h (P = 0.43) or 3 h (P = 0.10) after preparation. None of the labeling formulations differed significantly from the control values of the chemotactic index (all P > 0.25). For the complete blood count, no significant differences were observed between any of the groups, with the exception of platelet number in the control group (P = 0.004). The labeling efficiency (P = 0.10) and in vitro stability (P = 0.33) were also similar in our comparison of the 3 labeling formulations. Conclusion: With no major statistically significant difference noted either between the 3 labeling formulations or between the control and the 3 labeling formulations, we concluded that the methylene blue/phosphate buffer stabilizer (250 or 500 µg methylene blue) did not affect the cell membrane integrity, chemotactic capacity, labeling efficiency, in vitro stability, or complete blood count of leukocytes labeled with stabilized ^99mTc-exametazime.

Key Words: ^99mTc-exametazime • radiopharmaceutical stabilization • radiolabeled leukocytes • methylene blue • chemotaxis