全血法流式细胞术检测中性粒细胞抗体方法建立及临床初步应用

Establishment of the Method Detecting Antineutrophil Antibody in Whole Blood Sample With Flow Cytometry and its Primary Clinical Application

目的建立适用于临床的流式细胞术检测中性粒细胞(PMN)抗体的方法,确定健康人PMN CD16b和CD177两种抗原的参考范围,并初步用于检测抗PMN抗体。方法应用流式细胞术,以前向散射和侧向散射参数区分全血标本中的淋巴细胞和粒细胞,设定粒细胞门,用CD16b-PE和CD177-APC单克隆抗体分别检测中性粒细胞表面表达CD16b和CD177抗原的细胞的百分率,以此作为判断检测对象血液中抗CD16b或CD177抗体存在与否的依据。结果分别确定了健康人群PMN表达CD16b和CD177的参考范围(>x--2s)。心脑血管疾病患者3例中CD16b和CD177抗体均阳性者2例,淋巴瘤患者1例CD16b和CD177抗体均阳性,1例巨幼细胞性贫血患者检出CD16b抗体阳性,慢性PMN减少症患者6例中CD16b抗体阳性1例、CD16b和CD177抗体均阳性1例;自身免疫性疾病患者4例、甲状腺功能亢进、贫血患者和化疗后PMN减少的白血病患者各1例均未检出CD16b或CD177抗体阳性。结论流式细胞术检测抗中性粒细胞CD16b和CD177抗体的简便快捷,适用于自身免疫性中性粒细胞减少症的实验诊断和鉴别诊断。

Objective To establish the flow cytometry（FCM）method for clinical testing neutrophil antibodies,and determine the reference ranges of CD16band CD177 antigens on neutrophil of healthy control and primarily applicate it for testing neutrophil antibodies. Methods Application of flow cytometry,forward scattering and lateral scattering parameters to distinguish the whole blood samples of lymphocytes and granulocytes,to set the granulocyte gate,respectively percentage of neutrophil surface expression of CD16 b and CD177 antigen in cells with CD16b-PE and CD177-APC monoclonal antibodies to the object in the blood was selected to detect the anti CD16 b or anti CD177 antibody whether or not the basis. Results The reference ranges（ x--2s）of the expression of CD16 b and CD177 on neutrophil of healthy control were defined. There were both 2 cases of patients carried with anti-CD16 b and 3 cases patients carried with anti-CD177 antibodies,cardiovascular or cerebrovascular diseases and also both antiCD16 b and anti-CD177 antibodies in a patient with lymphoma,a patient with megaloblastic anemia,1 case of 6 patients with chronic neutropenia were checkouted anti-CD16 b antibodies,1 case of 6 patients with chronic neutropenia was checkouted both anti-CD16 b and anti-CD177 antibodies. There were no anti-CD16 b or anti-CD177 antibody in 4 patients with autoimmune diseases,1 patient with thyroid hyperfunction,1 patient with anemia and 1 patient with leukemia whose PMN decreased after chemotherapy. Conclusion The flow cytometry method for detecting anti-neutrophil CD16 b and CD177 antibody is simple and fast and well applicable to autoimmune neutropenia＇s laboratory diagnosis and differential diagnosis.