Dreier Jens, Vollmer Tanja, Kleesiek Knut
Institut für Laboratoriums- und Transfusionsmedizin, Herz- und Diabeteszentrum Nordrhein-Westfalen, Universitätsklinik der Ruhr-Universität Bochum, Bad Oeynhausen, Germany.
Clin Chem. 2009 Aug;55(8):1492-502. doi: 10.1373/clinchem.2008.122515. Epub 2009 Jun 4.
Bacterial contamination is the major infectious hazard associated with transfusion of platelet preparations (PLTs). Routine testing for bacterial contamination in PLTs has become common, but transfusion-transmitted bacterial sepsis has not been eliminated. Here, we describe a novel flow cytometry-based method for point-of-issue screening of PLTs for bacterial contamination.
We used the BactiFlow flow cytometer to detect and count bacteria based on esterase activity in viable cells. We compared the assay to incubation (BacT/Alert culture system) and rapid nucleic acid-based or immunoassay (reverse transcription PCR, Pan Genera Detection) methods.
We established a protocol for bacterial screening of PLTs consisting of enzymatic digestion and centrifugal filtration for the elimination of viable platelets and selective labeling of bacteria with fluorescent esterase substrate (ChemChrome V23). Results from the BactiFlow showed an excellent correlation (r = 0.9923 E. coli, r = 0.9736 S. epidermidis) to traditional plate count results. The lower detection limit of the assay was determined to be 150 CFU/mL, and the time to result was <1 h.
Our study demonstrates that BactiFlow flow cytometry is suitable for rapid screening of PLTs for bacterial contamination and fulfils the requirements for a point-of-issue testing of PLTs with acceptable time to result, specificity, sensitivity, and cost.
细菌污染是与血小板制剂（PLTs）输血相关的主要感染风险。对PLTs进行细菌污染的常规检测已很常见，但输血传播的细菌性败血症尚未消除。在此，我们描述了一种基于流式细胞术的新型方法，用于在发放点筛查PLTs的细菌污染情况。
我们使用BactiFlow流式细胞仪，根据活细胞中的酯酶活性来检测和计数细菌。我们将该检测方法与培养法（BacT/Alert培养系统）以及基于核酸的快速检测法或免疫检测法（逆转录PCR、泛属检测）进行了比较。
我们建立了一种用于PLTs细菌筛查的方案，该方案包括酶消化和离心过滤以去除活血小板，并用荧光酯酶底物（ChemChrome V23）对细菌进行选择性标记。BactiFlow的检测结果与传统平板计数结果显示出极佳的相关性（大肠杆菌r = 0.9923，表皮葡萄球菌r = 0.9736）。该检测方法的下限测定为150 CFU/mL，出结果时间<1小时。
我们的研究表明，BactiFlow流式细胞术适用于快速筛查PLTs的细菌污染情况，并且满足在可接受的出结果时间、特异性、敏感性和成本方面对PLTs发放点检测的要求。
