Lewis D C, McVey D S, Shuman W S, Muller W B
Department of Clinical Sciences, College of Veterinary Medicine, Kansas State University, Manhattan 66506, USA.
Am J Vet Res. 1995 Dec;56(12):1555-8.
To develop a flow cytometric assay for detection of platelet-bound IgG in dogs.
Negative-control platelets were obtained from 5 clinically normal Greyhounds. Positive-control platelets were platelets from 1 clinically normal dog, sensitized with dog anti-canine platelet alloantibodies.
Washed platelets were incubated with mouse anti-canine IgG conjugated to fluorescein isothiocyanate and analyzed by flow cytometry. Optimal dilution of antibody reagent and dose-response were determined, as were effects on platelet-bound IgG detection of storage time and temperature of K3EDTA-anticoagulated blood samples, variable platelet numbers, and variable filling of K3EDTA evacuated tubes.
A 1:128 dilution of antibody reagent was optimal. There was a linear increase in platelet-bound IgG when normal canine platelets were incubated with increasing concentrations of positive-control serum. Variable numbers of positive-control platelets tested and variable filling of K3EDTA evacuated tubes had no significant effect on platelet-bound IgG concentration. Platelet-bound IgG concentration increased with storage time at room temperature (P = 0.0003), but not when blood was kept cool. Sufficient platelets for assay were able to be isolated from 3 ml of blood from 5 dogs with < 10,000 platelets/microliters.
This assay for platelet-bound IgG in dogs is simple, repeatable, and practical. The assay is not affected by platelet count or variable filling of evacuated tubes, and requires only 3 ml of K3EDTA-anticoagulated blood. Blood samples for testing require packaging on ice and overnight delivery but, after arrival at the laboratory, can be refrigerated and analyzed within 72 hours of collection.
Assays for platelet-bound IgG may help in assessing causes and treatment of thrombocytopenia.
开发一种用于检测犬血小板结合IgG的流式细胞术检测方法。
阴性对照血小板取自5只临床正常的灵缇犬。阳性对照血小板来自1只临床正常的犬，用犬抗犬血小板同种抗体致敏。
将洗涤后的血小板与异硫氰酸荧光素偶联的小鼠抗犬IgG孵育，并用流式细胞术进行分析。确定了抗体试剂的最佳稀释度和剂量反应，以及K3EDTA抗凝血液样本的储存时间和温度、血小板数量变化和K3EDTA真空采血管填充量变化对血小板结合IgG检测的影响。
抗体试剂1:128稀释度最佳。当正常犬血小板与浓度递增的阳性对照血清孵育时，血小板结合IgG呈线性增加。检测的阳性对照血小板数量不同以及K3EDTA真空采血管填充量不同对血小板结合IgG浓度无显著影响。血小板结合IgG浓度随室温下储存时间的延长而增加（P = 0.（此处原文有误，应为0.0003）），但血液冷藏时则不会增加。对于血小板计数<10,000个/微升的5只犬，从3毫升血液中能够分离出足够用于检测的血小板。
这种犬血小板结合IgG检测方法简单、可重复且实用。该检测不受血小板计数或真空采血管填充量变化的影响，仅需3毫升K3EDTA抗凝血液。检测用血液样本需要冰上包装并隔夜送达，但到达实验室后可冷藏，并在采集后72小时内进行分析。
血小板结合IgG检测可能有助于评估血小板减少症的病因和治疗。
