Salgar S K, Paape M J, Alston-Mills B, Miller R H
Department of Animal Sciences, University of Maryland, College Park, MD 20742.
Am J Vet Res. 1991 Aug;52(8):1201-7.
A flow cytometric procedure was evaluated to measure the oxidative burst activity (hydrogen peroxide formation) of bovine neutrophils. The method involves measuring the oxidation of intracellular dichlorofluorescein to fluorescent dichlorofluorescein (DCF). Phorbol myristate acetate (PMA) was used to perturb the neutrophil plasma membrane. The sources of variation introduced into the DCF assay were also examined. The sources of variation were attributable to the isolation of neutrophils from blood, variation between duplicate assays and duplicate flow cytometric determinations of oxidative product formation, variation in neutrophil oxidative product formation among cows, and the variation (over repeated daily and weekly neutrophil isolations) in neutrophil oxidative product formation. A final objective was to determine effects of dexamethasone on oxidative product formation, and whether differences existed between blood and mammary neutrophils in oxidative product formation. There was an increasing trend in the formation of DCF with increasing time of incubation and with increasing PMA concentration. Increasing the concentration of PMA decreased lag time and increased the rate of oxidative product formation. The increase in DCF formation was statistically significant up to a PMA concentration of 10 ng/ml. This concentration was considered optimal for bovine neutrophils.(ABSTRACT TRUNCATED AT 250 WORDS)
对一种流式细胞术程序进行了评估，以测量牛中性粒细胞的氧化爆发活性（过氧化氢生成）。该方法包括测量细胞内二氯荧光素氧化为荧光二氯荧光素（DCF）。使用佛波酯（PMA）干扰中性粒细胞膜。还检查了DCF测定中引入的变异来源。变异来源可归因于从血液中分离中性粒细胞、重复测定和氧化产物形成的重复流式细胞术测定之间的变异、奶牛之间中性粒细胞氧化产物形成的变异以及（在每日和每周重复分离中性粒细胞过程中）中性粒细胞氧化产物形成的变异。最后一个目标是确定地塞米松对氧化产物形成的影响，以及血液中性粒细胞和乳腺中性粒细胞在氧化产物形成方面是否存在差异。随着孵育时间的增加和PMA浓度的增加，DCF的形成呈上升趋势。增加PMA浓度可缩短滞后时间并提高氧化产物形成速率。在PMA浓度达到10 ng/ml之前，DCF形成的增加具有统计学意义。该浓度被认为是牛中性粒细胞的最佳浓度。（摘要截断于250字）
