CE UPDATE- BLOOD BANKING II German F. Leparc, MD Leukocyte Reduction in Cellular Blood Components Downloaded from https://academic.oup.com/labmed/article/28/5/328/2503810 by guest on 27 September 2021 it is clear that some patients undergoing transfu­ ABSTRACT As we better understand the biologic effects of sion will benefit from the removal of leukocytes the presence of leukocytes in cellular blood components, we in their cellular blood components. must analyze the benefits, costs, and potential drawbacks ofleukodepletion procedures. Leukocyte removal can signifi­ Leukocyte Reduction Methods Methods for leukodepletion can be classified in cantly reduce the rate of febrile, nonhemolytic reactions as terms of removal rate in the following ways: well as the risk of cytomegalovirus infection. Leukodepletion • Low performance (<90%, 1 log reduction) can also prevent alloimmunization, bacterial growth, and • Intermediate performance (90%-99.9%, 1 to 3 storage lesions. We must address the timing of leukocyte log reduction) removal, quality assurance procedures, and specific indica­ • High performance (>99.9%) tions for each component to ensure that the appropriate Low-performance methods include saline wash, commitment of resources is made to leukocyte-reduction buffy coat removal, and spin-cool filtration. procedures. Intermediate-performance methods, such as glycerolization-freeze-thaw-deglycerolization This is the second article in a four-part series on blood banking. Other articles discuss new technologies in transfusion medicine, transfusion in the immuno­ (a method to remove leukocytes by repeated compromised host, and bone marrow transplantation. On completion of this series, washings), differential centrifugation, and the the reader will be able to list three advantages and disadvantages of the solid phase early adhesion-based filters (modified cotton technique, the gel test, and affinity column technology; recognize different wool or cellulose acetate), failed to consistently leukocyte reduction methods and identify the indications for the use of leukocyte- yield components with residual leukocyte counts reduced blood components; recommend strategies and select the most clinically less than 5 X 106. High-performance methods appropriate blood component therapy for immunosuppressed patients; and describe multiple advantages of DNA techniques compared with traditional include so-called third-generation filters that serologic methods for HLA typing. combine size retention, electrostatic attachment, and receptor-ligand interactions.6 When used From Florida Blood Leukocyte-reduced blood components (LRBCs) properly, these filters can deliver less than 6 Services, Tampa. are routinely available to physicians who order 5 X 10 leukocytes per unit transfused. Reprint requests to transfusions. As the biologic effects of collecting, In addition, manufacturers of apheresis Dr Leparc, Florida separating, storing, and transfusing blood or its equipment have developed platelet-harvesting Blood Services, 3602 components become better understood, the list Spectrum Blvd, technologies that allow platelets to be collected Tampa, FL 33612; of side effects attributed to the presence of leuko­ with minimal leukocyte contamination at levels ore-mail: gleparc@ cytes (white blood cells [WBCs]) continues to that consistently meet requirements for labeling as ix.netcom.com expand.1 Leukocytes have been perceived as an LRBC. potentially harmful to the viability of red blood cells2'3 and platelets2,4,5 intended for transfusion, Timing of Leukocyte Reduction and have been associated with adverse reactions Studies indicate that the timing of leukocyte in patients who are transfused (Table 1). To removal may be critical in achieving benefits confuse matters further, leukocytes may have linked with WBC depletion. During storage, the beneficial effects based on empiric observations release of cytokines (particularly in platelet com­ that have not been proven (Table 2). Nevertheless, ponents, because the leukocytes are metabolically active at the higher storage temperatures that platelets require) and proteases (responsible in Downloaded from https://academic.oup.com/labmed/article-abstract/28/5/328/2503810 by guest LABORATORY MEDICINE VOLUME 28, NUMBER 5 MAY 1997 on 28 May 2018 part for the structural changes in cellular ele­ benefits of leukodepletion in the treatment of ments that are described generically as "storage certain patients. It does not appear necessary to lesions") begins within a few hours of collection.7 remove leukocytes in routine blood processing in By removing leukocytes within the first 24 hours view of the higher costs involved. of collection, the source of the noxious leukocytic Although no single indication has yielded suf­ byproducts is removed before significant levels ficient data to qualify as a requirement for leuko- build and cause measurable effects. reduction, some generally accepted indications Although no controlled human studies have for the use of LRBCs follow: been published supporting the claim, animal • Prevention of febrile, nonhemolytic transfusion models support the proposed theoretic benefits Downloaded from https://academic.oup.com/labmed/article/28/5/328/2503810 by guest on 27 September 2021 reactions, particularly in multitransfused patients of early leukodepletion in the prevention of who have experienced this complication in past alloimmunization to HLA.8'9 transfusions. Taking into consideration that Moreover, some studies suggest that early removal of leukocytes may significantly decrease bacterial contamination in stored units.10"13 Language of Leukodepletion Bacteria that enter the unit at collection time are phagocytized (ingested) by leukocytes soon after­ Leukocyte—A white blood cell or corpuscle ward. The dangers of contamination are averted by removing the bacteria-laden cells within 24 Leukocyte load—The amount of white blood cells present in the hours of collection. component to be processed. It is probably the most important element in the leukoreduction equation, because different leukocyte Quality Control Procedures depletion methods vary in their load capacity. On average, a unit Each institution must validate its leukocyte- of whole blood contains from 2 to 3 X 109 white blood cells. reduction procedures to ensure that the equip­ Components collected by apheresis procedures vary in their ment and method in use consistently meet the leukocyte load depending on the instrument, collection settings, residual-leukocyte counts and product-recovery and donor white blood cell count. The final product may contain rates claimed. Laboratory or bedside filters must from less than 1 X 106 to more than 5 X 109. be validated each time a new method or filtering device is implemented.14 A sampling plan should Leukodepletion—The process of removing white blood cells be developed to provide a level of confidence that Leukoreduction rate—The proportion of the total leukocyte load errors in the application of leukocyte-reduction that is effectively removed from the original component. It may be procedures are detected and corrective action is expressed as a percentage (eg, 99.5%) or as a logarithmic implemented. expression (eg, 3 logs). The two terms may be used interchangeably, Manual or automated procedures for counting and log reductions (ie, base 10 logarithmic expressions of leukore­ the number of residual WBCs must be estab­ 0 duction rates) can be converted readily to percentages as follows: U lished. The method in use must meet sensitivity u thresholds and linearity requirements to accu­ 1 log reduction = 90% rately count small numbers of leukocytes 2 log reduction = 99% 14 0 (<3 WBC/|xL) in the processed components. 3 log reduction = 99.9% H Methods for counting WBCs in very low con­ 4 log reduction = 99.99%, and so on centrations include the following: Recovery rate—The amount (expressed as a percentage) of cells • Manual counts using the Nageotte (Bright Line, intended for transfusion in the original component that are left u Hauser Scientific, Horsham, Pa) chamber (done after the leukocyte reduction process is completed. The recovery rate either on fresh samples or aliquots fixed with 3% should be as high as possible to maximize the benefits of transfusion. 15 paraformaldehyde for batch testing) Recommendations issued by the Food and Drug Administration's • Low cytometry (conventional or automated Center for Biologies Evaluation and Research (CBER)* state that volumetric capillary cytometry16) methods used must "retain a minimum of 85% of the original • Polymerase chain reaction product." Residual leukocyte count—The amount of white blood cells found in Indications for the Use of the component after leukoreduction. CBER recommendations state Leukocyte-Reduced Blood Components that cellular blood components labeled with the "leukocytes reduced" Consensus exists regarding the benefits of remov­ modifier must be prepared "by a method known to leave a residual ing "passenger" leukocytes in cellular blood leukocyte count of less than 5 X 106." components as well as the proven and potential •FDA memo.18 Downloaded from https://academic.oup.com/labmed/article-abstract/28/5/328/2503810 MAY 1997 VOLUME 28, NUMBER 5 LABORATORY MEDICINE 329 by guest on 28 May 2018 TABLE 1. ADVERSE REACTIONS ASSOCIATED WITH THE PRESENCE OF DONOR LEUKOCYTES residual leukocyte counts when used for this pur­ 6 Febrile, nonhemolytic transfusion reaction pose also should be less than 5 X 10 . Alloimmunization to leukocyte antigens