多参数流式细胞术分析急性白血病免疫表型

为了探讨急性白血病免疫表型特征,利用流式细胞术三色荧光直接标记技术CD45/SSC多参数散点图设门方法,对162例急性白血病患者幼稚细胞表面及胞浆内分化抗原进行分析.结果提示:急性非淋巴细胞白血病(ANLL)患者主要表达CD117(94.9%)、CD13(88.5%)和CD33(70.5%);急性淋巴细胞白血病(ALL)患者中B-ALL主要表达cCD79a(100%)、CD19(92.1%),T-ALL表达cCD3(100%)、CD2(83.3%);ANLL伴淋系抗原表达(Ly ANLL),以CD7(56.2%)、CD19(31.2%)多见;ALL伴髓系抗原表达(My ALL),以CD13(88.9%)、CD33(27.8%)多见.为此用多参数流式细胞术三色荧光直接标记法进行免疫分型,对急性白血病的确诊、治疗和预后有重要的临床意义.     

流式细胞术在我院急性白血病免疫分型中的应用

目的探讨本实验室所用13种单抗对白血病免疫分型的应用价值。方法应用流式细胞术（FCM）使用13种单抗对99例急性白血病进行免疫分型,与形态学分型相比较。结果使用13种单抗从99例急性白血病中分出髓系白血病（AML）59例、淋系白血病（ALL）27例、混合型（MAL）5例、未分化型（AUL）2例,诊断率达93.9%（93/99）。AML中各抗原表达情况：CD13〉CD33〉CD14、CD7〉CD2、CD19〉CD10〉CD5〉CD20,其中多表达CD13、CD33;ALL中各抗原表达情况：CD19〉CD13〉CD10〉CD20〉CD33〉CD5〉CD7〉CD2、CD14,其中CD10、CD13、CD19、CD20表达率较高。免疫分型与形态学分型结果相比较,AML二者的吻合率为90.5%（57/63）、ALL二者的吻合率为76.5%（26/34）;其中1例M0免疫分型为B/M混合、1例M1免疫分型为T/M混合、1例M2a免疫分型为T/M混合、1例L1免疫分型为B/M混合、1例L2免疫分型为T/B混合、1例L2免疫分型为Ly＋-AML、2例L2免疫分型为AUL;1例形态学为浆细胞白血病的免疫分型为AML、1例淋单混合型白血病CD7、CD13、CD14阳性。结论白血病细胞、免疫分型各具特征,形态学相同的白血病免疫分型未必相同,形态学不同的白血病免疫分型未必不同;本实验室所选单抗组合基本能满足白血病的免疫分型,对于一些混合型和未分化型白血病的诊断亦能明确。     

多色流式细胞术对40例急性白血病免疫分型的研究

白血病的形态学、免疫学、细胞遗传学和分子生物学分型已成为现代白血病诊断所必需,其中流式细胞术免疫分型已成为急性白血病(AL)诊断和分型的重要依据,具有重要的临床诊断意义.我们采用流式细胞仪四色荧光标记技术,CD45/SSC双参数散点图设门方法,可全面地与特异地检测出白血病细胞抗原的分布,使白血病免疫分型的结果更为准确和客观[1].     

急性髓系白血病177例免疫分型特点及预后分析

目的:探讨急性髓系白血病(AML)患者免疫分型的特点及临床意义.方法:采用CD45/SSC双参数散点图设门方法对177例急性髓系白血病患者进行三或四色流式细胞术免疫分型分析.结果:177例AML显示,CD33,CD38,CD117,CD13及HLA-DR高度表达,阳性率分别为92.66%,81.36%,75.14%,68.93%和67.80%.40.7%的AML患者伴有淋系抗原表达,最常见的是CD7(22.6%),其次是CD19(7.91%).结论:多色流式细胞仪术免疫分型对AML的诊断及预后具有重要意义.     

141例白血病三色流式细胞术免疫分型研究

白血病是常见的恶性肿瘤之一，其分型80年代以前主要是依靠形态学，自单克隆抗体与流式细胞术联合应用以来，使诊断白血病的准确率提高到90％以上，从而为临床合理治疗、判断预后提供了重要的科学依据。为了克服流式细胞术单参数免疫标记测定易将正常细胞包括在内从而影响结果分析的不足，我们采用以CD45和侧向角散射（side scatter，SSC）双参数散点图设门，应用三色流式细胞术对141例白血病患者进行免疫分型研究，现将结果报道如下。     

195例急性白血病免疫表型分析

应用流式细胞技术可快速确定白血病细胞胞膜或胞质内的抗原,通过分析自血病细胞的抗原表达,来了解白血病细胞所属细胞系列及其分化程度,这就是白血病免疫分型。本文通过流式细胞术对195例急性白血病（AL）的免疫表型进行了分析。     

多参数流式细胞术对外周血白血病免疫分型的研究

目的 :探讨多参数流式细胞术、CD45/SCC设门技术和三色荧光标记抗体 ,检测外周血在白血病免疫分型中的临床应用及其意义。方法 :对检测标本以CD45- Percp/SSC设门技术及流式细胞仪多参数分析技术 ,采用单克隆抗体三色标记对白血病患者的细胞群体进行免疫表型分析。结果 :195例白血病患者 ,其中急性白血病 14 4例 ,慢性白血病 51例。 14 4例急性白血病患者中 ,13 7例出现可供分析的白血病细胞独立群体。免疫分型结果ALL 66例 ,AML 67例 ,未能分型 4例。 2 7例AL表达 2个不同系列的免疫标记 (2 0 % )。分析了各类白血病的免疫表型的特征 ,AL共表达不同系列的抗原是一种常见现象。结论 :利用CD45 Percp/SSC设门技术及流式细胞仪多参数分析技术 ,可对白血病细胞群体进行准确的免疫表型检测和研究 ,有助于指导临床治疗及判断预后     

免疫表型与急性白血病的分型诊断

对426例急性白血病采用APAAP法23种单抗进行免疫分型研究.结果发现:(1)426例中,71.6%可确定为淋巴系或髓系或髓/淋混合型,28.4%不能确定细胞系列.FAB结果仅62.75%得到免疫表型证实,29.5%不能判断细胞起源,7.75%两者分型结果不一致;(2)ANLL140例中,44.29%免疫表型证实为髓细胞型,17.14%仅表达淋巴系标志,38.57%不能判断细胞起源,各髓系标志无明显FAB亚型特异性;(3)ALL260例中,72.67%进一步证实为淋巴细胞型,2. 69%单纯表达髓系标志或混合表达髓系和淋巴系抗原,另外24.62%不能确定细胞起源;(4)FAB不能分型26例中,23例可确定为ALL或ANLL,另外3例仍不能诊断.研究认为,免疫分型是FAB分型的必要补充.     

急性白血病患者骨髓细胞形态和免疫表型同步检测的比较分析

目的 探讨骨髓细胞免疫表型在急性双表型白血病中的诊断价值以及与骨髓细胞形态学的比对研究.方法 利用流式细胞仪四色免疫荧光直接标记技术对230例急性白血病患者进行细胞免疫表型、骨髓细胞形态学分型检测及分析.采用流式细胞术四色免疫荧光直接标记技术与CD45/SSC设门分析技术检测13例双表型急性白血病患者,根据欧洲白血病免疫分类组(EGIL)积分标准进行免疫学分型.结果 230例急性白血病患者免疫分型诊断结果与骨髓细胞形态学和组织化学诊断具有高符合率,其中免疫分型诊断为急性双表型白血病的13例(5.7％).此外,急性双表型白血病患者高表达CD34胞膜抗原(84.6％),提示预后不良.结论 急性双表型白血病发病率较低,骨髓细胞免疫表型对诊断及鉴别双表型急性白血病极为特异,以髓系和B淋巴系抗原共表达为主.应用骨髓细胞形态学和流式细胞术联合检测急性双表型白血病可以提高诊断的准确性,有效地指导临床制定治疗方案和预后判断.     

白血病免疫分型诊断技术的进展

白血病是造血系统的恶性疾病,俗称“血癌”,是国内十大高发恶性肿瘤之一.目前,免疫分型已是当今白血病诊断分型、预后判断和残留病检测的一个有力手段.免疫分型诊断的主要方法有免疫细胞化学技术、流式细胞术、细胞芯片技术,也有个别学者在尝试压电免疫传感器.临床上仍以流式细胞术(FCM)方法为主,目前一次最多能检测到15种抗原,且昂贵.而细胞免疫芯片检测技术,一次可以检测数十种乃至上百种细胞抗原,如能应用于临床,将是一种方便,快速,简单,经济实用的白血病免疫分型检测手段.     

急性髓细胞白血病患者CD56抗原表达与MDR1基因表达量的关系研究

目的 研究初治急性髓细胞白血病(AML)患者CD56抗原表达与多药耐药(MDR)1基因表达量的关系,探讨CD56抗原表达与MDR1基因表达量在AML耐药中的作用及相互关系.方法 采用流式细胞术(FCM)和建立实时荧光定量PCR技术分别检测79例AML患者CD56抗原表达及MDR1基因表达水平并分析两者之间的关系及临床意义.结果 24.1%AML患者表达CD56抗原,FAB亚型中M5 AML患者表达阳性率高于其他亚型.遗传学危险度分级高危组患者CD56抗原表达阳性率显著高于中危组(P<0.01),伴t(8:21)AML患者CD56抗原表达阳性率(57.1%)显著高于其他低危组AML(P<0.05).CD56抗原表达阳性初治AML患者MDR1基因表达水平显著高于表达阴性患者(P<0.001).MDR1基因高表达且CD56抗原阳性AML组的CR率(58.8%)显著低于MDR1基因低表达且CD56表达阴性组(89.2%,P<0.01).结论 AML患者CD56抗原表达与MDR1基因表达水平存在相关性;同时定量检测MDR1基因表达及CD56抗原表达更有助于判断预后.     

急性淋巴细胞性白血病微小残留病变筛选指标的特点分析

目的 对儿童急性淋巴细胞性白血病微小残留病变(MRD)筛选指标进行分析并评估其意义和表达特点.方法 分离35例初发B-急性淋巴细胞性白血病(ALL)患儿的单个核细胞,对符合CD38、CD45弱表达,CD58、CD21、CD22强表达,CD34和Cu同时表达,染色体相关抗原CD66c表达的,与CD10/CD34/CD19进行四色抗体组合,应用流式仪检测,如在双参数点图上所选择的四色抗体组合出现的位置明显有别于正常骨髓相应位置的,则认定该抗体组合为有效的筛选标记并进行随后的MRD监测.结果 35例患儿中31例存在至少一个MRD标记,覆盖率为88.6%;21/35例患儿(60%)存在2个或2个以上的筛选标记;TdT/CD10/CD34/CD19为最常见的四色组合.结论 TdT/CD10/CD34/CD19作为四色MRD筛选标记覆盖率高,应作为常规和首选的筛选标记;免疫表型中Pro-B缺乏有效的筛选标记,出现2个或以上的筛选标记,对提高MRD的精确度具有重要意义.     

Role of flow cytometry in diagnostics of myelodysplastic syndromes--beyond the WHO 2008 classification

Multiparameter flow cytometry (FCM) is an excellent method to follow the expression patterns of differentiation antigens using monoclonal antibodies to surface and cytoplasmic proteins. Although several authors described various aberrant immunophenotypic features in the bone marrow of patients with myelodysplastic syndromes (MDS), the World Health Organization 2008 classification recommended that, only if 3 or more phenotypic abnormalities are found involving 1 or more of the myeloid lineages can the aberrant FCM findings be considered suggestive of MDS. In the absence of conclusive morphologic and/or cytogenetic features, FCM abnormalities alone were considered not sufficient to establish MDS diagnosis and further follow-up of the patients was recommended. Review of the literature gives accumulating evidence that FCM has become an important part of the integrated diagnostic work-up of patients with suspected MDS. Several studies have also reported FCM findings significant for prognosis and therapy choice in MDS patients. Technical progress in multicolor FCM and new analysis programs, together with ongoing efforts to standardize the methodology, will make it possible to apply FCM in individual risk assessment and choice of best therapy for MDS patients.     

Immunophenotypic, cytogenetic and clinical features of 192 AML patients in China

Immunophenotypic characterization of the leukemic cells has been widely used as a tool for diagnosis, classification and prognosis of leukaemia. A total of 192 Chinese patients with acute myeloid leukemia (AML) were immunophenotyped by flow cytometry using a panel of monoclonal antibodies. Among the 192 patients enrolled in this study, 125 cases were also subjected to karyotype analysis by G-banding technology. We found that CD33, CD13, MPO and CD117 were the most commonly expressed antigens in AML. A combination of intensive autofluorescence, both CD34⁻ and HLA-DR⁻, and high expression of CD13, CD33 and MPO had significant value for M3 diagnosis. CD14 was expressed only in M4 and M5, and both intensive positivity of CD64 and CD15 with high expression of HLA-DR may suggest great possibility for diagnosis of M5. Lymphoid markers expression was documented in 47.9% of the 192 AML cases analyzed. CD56 (26.0%) and CD7 (20.8%) were the most commonly expressed lymphoid markers in AML patients. Abnormal karyotypes were detected in 76 out of 125 (60.8%). Higher CD34 positivity was found in LymAg⁺ group (77.2%) than in LymAg⁻ group (48.0%). Our results indicate that immunophenotype analysis was useful for AML diagnosis and classification and the immunophenotype did have relevance to the abnormal cytogenetic changes and clinical features in AML.     

Associations of killer cell immunoglobulin-like receptors with acute myeloid leukemia in Chinese populations

Many studies have investigated the relationship between KIR, HLA and acute myeloid leukemia (AML), but the results were different in different laboratories, and the data in Chinese population were limited. In this study, the distribution of KIR gene, KIR genotypes, HLA-C groups, HLA-Bw4, and KIR-HLA interaction from 273 healthy participants and 253 AML patients (M0–M6) in southern Chinese Han were determined to investigate the relationships among KIR, HLA and AML. The results showed that the frequencies of 2DS4del in M5 patients were significantly higher than those of the controls (65.0% vs 46.5%, P = 0.0104, OR = 2.135, P < ɑ′). The frequency of KIR genotype BX13 in the healthy controls was significantly higher than that in AML patients (3.7% vs 0%, P = 0.0019, OR = 20.2, P < ɑ′). No other significant differences in the frequencies of KIR, HLA and KIR-HLA interaction were identified between AML patients and controls. Our study suggests that 2DS4del may conduct a susceptibility to AML, and genotype BX13 might conduct a protective effect on AML.     

急性早幼粒细胞白血病68例回顾性临床分析

目的探讨初治中低危急性早幼粒细胞白血病(APL)患者诱导缓解治疗及完全缓解(CR)后的治疗方案。方法回顾性分析68例初治APL患者3种不同诱导治疗方法CR率、达CR所需时间及副作用,并分析CR后不同巩固治疗方案对预后的影响。结果全返式维甲酸(ATRA)联合蒽环类药物化疗、单用亚砷酸(AS2O3)及ATRA+AS2O3+化疗3种方法治疗初诊APL完全缓解率分别为90%、89%、89%,无显著差异,P>0.05;ATRA+AS2O3+化疗组达CR所需时间最短,ATRA+化疗组副作用最少,P<0.05;CR后选用ATRA、AS2O3、化疗序贯治疗能明显延长患者无病生存期。结论 ATRA+化疗方案可做为初治APL患者诱导缓解的首选方案;ATRA、AS2O3、化疗序贯治疗均可做为CR后的有效治疗方案,中低危患者可不必联用Ara-c化疗。     

Comprehensive flow cytometry phenotype in acute leukemia at diagnosis and at relapse

Li X, Du W, Liu W, Li X, Li H, Huang S‐A. Comprehensive flow cytometry phenotype in acute leukemia at diagnostic and at relapse. APMIS 2010; 118: 353–59. Multiparameter flow cytometry (MFC) plays a vital role in the detection of minimal residual disease (MRD) and diagnosis of relapse in acute leukemia. However, application of a limited panel of antibodies in MFC leads to high rates of false‐negative and false‐positive results. Thirteen patients with acute lymphoblastic leukemia (ALL) and 12 patients with acute myeloid leukemia (AML) were immunophenotyped by MFC at diagnosis and at relapse using a comprehensive panel of monoclonal antibodies (McAbs) to 27 antigens and CD45/SSC gating. In 23 of 25 patients (92.3%), changes in at least one of progenitor‐associated, myeloid and lymphoid antigens between diagnosis and relapse were observed. Antigen changes were observed in 92 of 239 antigens (38.5%) expressed in 25 patients, in 49 of 117 antigens (41.9%) expressed in 13 ALL patients, and in 43 of 122 antigens (35.2%) expressed in 12 AML patients. Phenotypic changes were characterized by the expression of cross‐lineage antigens. The intralineage change was observed in the majority of patients. However, myeloid lineage shift was identified by MFC in two patients with T‐ALL. Multiple panels of three or more McAbs are likely to be required in the monitoring of MRD and diagnosis of relapse in acute leukemia by MFC.