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Updated: Sep 16, 2026
Bioenergetics and the Oxidative Burst: Protocols for the Isolation and Evaluation of Human Leukocytes and Platelets
Published on: March 27, 2014
Function of irradiated polymorphonuclear leukocytes obtained by buffy-coat centrifugation
Abstract:
Several studies suggest that transfusion of polymorphonuclear leukocytes (PMNs) may be beneficial in the treatment of septic neonatal patients. Because of expense, donor availability, and the technical effort involved in obtaining PMNs by intermittent or continuous flow leukapheresis, buffy coat centrifugation of whole blood has been suggested as an alternative source. An in vitro study was performed to determine whether PMNs collected by this method have adequate oxidative and migratory function measured by chemiluminescence (CL) and chemotaxis under agarose (CT), respectively. Whole blood samples from six adult volunteers were drawn into citrate-phosphate-dextrose-adenine-one and stored at 4 degrees C for 0 to 48 hours. One-half of each sample was irradiated with 1500 rads. PMNs isolated from the buffy coat of these samples had greater than 80 percent normal CT and CL following 0 to 28 hours of storage in whole blood. Irradiation caused no depression in function. Units of whole blood yielded 1.11 +/- 0.40 X 10(9) PMNs per unit. This study indicates that transfusion of radiated PMNs obtained from stored whole blood that is less than 28 hours old is reasonable to use in studies involving PMN transfusions.
Insights
Buffy coat centrifugation is a viable method for obtaining polymorphonuclear leukocytes (PMNs) for neonatal sepsis treatment. Irradiated PMNs from stored whole blood maintain adequate function for transfusion studies.
Area of Science:
- Hematology
- Immunology
- Neonatal Medicine
Background:
- Polymorphonuclear leukocytes (PMNs) transfusions show promise for treating neonatal sepsis.
- Current methods for PMN collection (leukapheresis) are costly and complex.
- Buffy coat centrifugation is proposed as a more accessible alternative.
Purpose of the Study:
- To evaluate the oxidative and migratory functions of PMNs isolated via buffy coat centrifugation.
- To assess the impact of storage and irradiation on PMN function.
Main Methods:
- Whole blood samples from six volunteers were collected and stored at 4°C for up to 48 hours.
- Half of each sample was irradiated (1500 rads).
- PMN function was assessed using chemiluminescence (CL) and chemotaxis under agarose (CT).
Main Results:
- PMNs retained over 80% of normal CT and CL function after 0–28 hours of storage.
- Irradiation did not significantly impair PMN function.
- One unit of whole blood yielded approximately 1.11 x 10^9 PMNs.
Conclusions:
- Buffy coat centrifugation is a practical method for obtaining functional PMNs.
- Irradiated PMNs from stored whole blood (less than 28 hours old) are suitable for transfusion studies.
- This method offers a cost-effective alternative for PMN procurement in neonatal care.
