de Back Djuna Z, Vlaar Richard, Beuger Boukje, Daal Brunette, Lagerberg Johan, Vlaar Alexander P J, de Korte Dirk, van Kraaij Marian, van Bruggen Robin
Department of Blood Cell Research, Sanquin Research, and Landsteiner Laboratory, University of Amsterdam, Amsterdam, The Netherlands.
Department of Product and Process Development, Sanquin Blood Bank, Amsterdam, The Netherlands.
Transfusion. 2018 Apr;58(4):896-904. doi: 10.1111/trf.14535. Epub 2018 Feb 15.
Several circumstances require the accurate measurement of red blood cell (RBC) survival and clearance, such as determination of posttransfusion recovery of stored RBCs to investigate the effect of new additive solutions. To this end, biotin as a marker of RBCs to track donor RBCs in the blood of the recipient has been used in many studies. However, so far only experimental, nonvalidated, biotin-labeled red cell concentrates (RCCs) are transfused. The goal of this study was to produce a standardized biotin-labeled RCC product in a fast, simple, and sterile manner that can be used for clinical research and for the evaluation of new blood products according to Good Practice Guidelines (GPG) for blood establishments.
RCC fractions were labeled with two different concentrations of biotinylation reagent in a closed system, to prevent bacterial contamination of the end product. Using flow cytometry, the reproducibility and robustness of the biotin labeling was assessed, as well as the stability of the biotin label on the (un-)irradiated RCC fraction. Additionally, parameters such as phosphatidylserine (PS) exposure, sodium (Na), potassium (K), free hemoglobin, adenosine triphosphate (ATP), pH, and morphology were determined prior to and after biotin labeling to rule out detrimental effects of the labeling procedure on the RCC.
Our data show that RCCs can be labeled under sterile conditions in a closed system with two different biotinylation reagent concentrations, without affecting the biological activity.
An easy, rapid (<2 hr), and robust method was developed to manufacture biotin-labeled RCCs for clinical research compliant to GPG.
在几种情况下需要准确测量红细胞（RBC）的存活和清除情况，例如测定储存红细胞的输血后恢复情况以研究新型添加剂溶液的效果。为此，许多研究使用生物素作为红细胞标记物来追踪受者血液中的供体红细胞。然而，到目前为止，仅输注了实验性的、未经验证的生物素标记红细胞浓缩物（RCC）。本研究的目的是以快速、简单和无菌的方式生产标准化的生物素标记RCC产品，该产品可用于临床研究以及根据血液机构的良好操作规范（GPG）评估新型血液制品。
在封闭系统中用两种不同浓度的生物素化试剂标记RCC组分，以防止最终产品受到细菌污染。使用流式细胞术评估生物素标记的可重复性和稳健性，以及生物素标记在（未）辐照RCC组分上的稳定性。此外，在生物素标记前后测定诸如磷脂酰丝氨酸（PS）暴露、钠（Na）、钾（K）、游离血红蛋白、三磷酸腺苷（ATP）、pH和形态等参数，以排除标记过程对RCC的有害影响。
我们的数据表明，RCC可以在封闭系统中在无菌条件下用两种不同浓度的生物素化试剂进行标记，而不影响其生物活性。
开发了一种简便、快速（<2小时）且稳健的方法来生产符合GPG的用于临床研究的生物素标记RCC。
