Morikawa K
Rinsho Byori. 1989 Sep;37(9):1013-9.
To examine granulocyte oxidative metabolism (GOM) quantitatively, we evaluated two methods, one for assaying luminol-dependent chemiluminescence (CL) and the other for changes in fluorescence by dichlorofluorescin (DCF). The CL assay was carried out using a lumiphotometer (TD-4000, Laboscience) after stimulating granulocytes in whole blood by n-formyl-methonyl-leucyl-phenylalanine (FMLP, 100 nmol/l) or by opsonized zymosan (OZ, 2 mg/ml). To reduce hemoglobin (Hb) interference, each sample was diluted with autologous plasma to a Hb concentration of 3 g/dl. The DCF assay was performed on whole blood samples (100 microliters) labeled with DCF (5 mumol/l) following stimulation by phorbol myristate acetate (PMA, 100 ng/ml). Changes in fluorescence were determined using a flow cytometer that gave the delta mean channel fluorescence intensity (DMCF) as an indicator of GOM. The coefficients of variance (CV) in within-run assays for the CL and the DCF were 10% and 5%, respectively. These CV-values were almost the same even when separated granulocytes as a test sample were used instead of whole blood. CL determined by FMLP stimulation in 27 renal failure patients on hemodialysis (HD), was significantly lower than that in 10 normal controls, but no difference was found in that determined by OZ stimulation. In HD patients, the DMCF values tended to increase in those at 15 min and the end of HD sessions compared to the value prior to HD. This suggested that the HD membrane and/or extracorporeal circulation stimulates GOM.(ABSTRACT TRUNCATED AT 250 WORDS)
为了定量检测粒细胞氧化代谢（GOM），我们评估了两种方法，一种用于检测鲁米诺依赖性化学发光（CL），另一种用于检测二氯荧光素（DCF）引起的荧光变化。CL检测是在通过N-甲酰甲硫氨酰亮氨酰苯丙氨酸（FMLP，100 nmol/l）或调理酵母聚糖（OZ，2 mg/ml）刺激全血中的粒细胞后，使用发光光度计（TD-4000，Laboscience）进行的。为了减少血红蛋白（Hb）的干扰，每个样本用自体血浆稀释至Hb浓度为3 g/dl。DCF检测是在佛波酯（PMA，100 ng/ml）刺激后，对用DCF（5 μmol/l）标记的全血样本（100微升）进行的。使用流式细胞仪测定荧光变化，以平均通道荧光强度变化（DMCF）作为GOM的指标。CL和DCF的批内检测变异系数（CV）分别为10%和5%。即使使用分离的粒细胞作为测试样本代替全血，这些CV值也几乎相同。27例接受血液透析（HD）的肾衰竭患者经FMLP刺激测定的CL显著低于10例正常对照，但经OZ刺激测定的CL无差异。在HD患者中，与HD前的值相比，15分钟时和HD结束时的DMCF值往往会增加。这表明HD膜和/或体外循环刺激了GOM。（摘要截断于250字）
