免疫双标记细胞的检测在中枢神经系统白血病诊断中的意义

目的：观察末端脱氧核苷酸转移酶（TdT）及淋巴细胞系列相关抗原双标记细胞在脑脊液中的检出对中枢神经系统白血病（CNS－L）的临床意义。方法：采用免疫荧光标记技术配合流式细胞术，检测初治或安全缓解期急性淋巴细胞白血病脑脊液中TdT及淋巴系列相关抗原双标记细胞及其治疗后的变化。结果：64例中8例脑脊液检出上述免疫双标记细胞，均为急性淋巴细胞白血病，其中1例无CNS－L临床迹象，免疫双标记细胞占3.38%，经治疗后免疫双标记细胞在脑脊液均明显减少或消失。结论：脑脊液中免疫双标记细胞的检出对CNS－L的诊断及治疗指导具有重要的临床意义。弥补了常规细胞形态学的不足。     

91例急性白血病FCM免疫分型分析

目的：研究急性白血病免疫分型及临床意义。方法：采用流式细胞术检测91例急性白血病患者免疫分型情况。结果：①80％以上急性非淋巴细胞白血病(ANLL)患者主要表达CD13、CD33。②按免疫表型的特征可将急性白血病分为3类：系列专一性表达；交叉表达；“裸细胞”型。ANLL中，以系列专一性比例最高，交叉表达在ALL中占有一定比例，“裸细胞”型在ANLL和ALL中比例均最少。交叉表达的病例中，CD7^ ANLL患者CR率低于系列专一性表达者。结论：AL免疫分型可出现系列专一性表达；交叉表达；“裸细胞”型3种类型，CD7^ ANLL交叉表达的患者完全缓解率低于系列专一性表达者。     

流式细胞术检测双表型急性白血病

目的：应用流式细胞术检测双表型急性白血病。方法：用直接荧光标记的单克隆抗体分析双表型急性白血病的免疫表型。结果：①双表型白血病的构成比例是3．16％（28／886）,其中B／M双表型白血病占64．28％,T／M双表型白血病占25．。0％,T／B双表型白血病占10．71％。②B／M双表型白血病均共同表达细胞质cCD79a和cMPO;T／M双表型白血病均共同表达细胞质cCD3和cMPO;T／B双表型白血病均共同表达细胞质cCD3和cCD79a。③形态学诊断为急性非淋巴细胞白血病4例,急性淋巴细胞白血病6例,经流式细胞术分析均为双表型急性白血病。结论：①双表型白血病以B／M系抗原共同表达最常见;②cCD3、cCD79a、cMPO为系列特异性标志抗原,对诊断及鉴别双表型急性白血病极为重要;③流式细胞术是目前诊断双表型急性白血病特异可靠的方法。     

白血病免疫表型分析

目的：应用流式细胞术研究白血病免疫表型的特点.方法：采用单克隆抗体直接免疫荧光标记法，对209例白血病患者进行免疫表型分析.结果：CD13、CD33是急性髓细胞白血病特异和敏感的指标，CD2、CD7是T-淋巴细胞白血病（T-ALL）特异和敏感的指标，CD19、CD10是B-ALL特异和敏感的指标.结论：运用流式细胞术对白血病进行免疫分型可以极大提高诊断的准确性，指导临床治疗和判断预后。     

c-kit受体与其他髓系膜抗原标志物对急性非淋巴细胞白血病诊断价值的比较

近年来，有报道称c-kit受体(c-kitR，CD117)可作为髓系免疫学标记物，用于急性白血病(AL)的分型诊断，对急性非淋巴细胞白血病(ANLL)的诊断以及ANLL与急性淋巴细胞白血病(ALL)的鉴别诊断有重要参考价值。本研究运用流式细胞术(FCM)检测CD117在AL中的表达，探讨其对ANLL的诊断意义；并根据灵敏度和特异度的概念将CD117与CD13、CD14、CD33等髓系膜抗原标志物对ANLL的诊断价值进行了比较研究。     

髓系抗原在成人急性淋巴细胞白血病中的临床意义

髓系抗原在成人急性淋巴细胞白血病中的临床意义李建勇夏学鸣唐晓文郑列林薛永权阮长耿免疫分型对于急性淋巴细胞白血病（急淋）的诊断、分型及预后判断具有重要作用［１］。为了解免疫分型，特别是髓系抗原表达（Ｍｙ＋）在成人急淋中的临床意义，对６１例初治患者进行了...     

电镜及免疫标记髓过氧化物酶检测对急性白血病分型诊断的意义

目的 比较电镜髓过氧化物酶(MPO)、免疫标记抗髓过氧化物酶抗体(抗MPO抗体)、光镜过氧化物酶(POX)对急性白血病分型诊断的敏感性和特异性。方法 对38例初诊的急性白血病患者，APAAP法检测MPO抗体及电镜MPO细胞化学染色。结果 光镜POX阳性组抗MPO及电镜MPO染色均呈阳性，光镜POX阴性组22例中有5例抗MPO抗体阳性，6例电镜MPO染色阳性。结论 抗MPO法及电镜化染对急性白血病分类中髓系特征的判断具有敏感性好，特异性高的优点，个别病例电镜MPO反应较免疫标记MPO抗体更为有效和敏感。     

CD117和CD34在急性白血病中的临床意义

目的:观察细胞表面分化抗原CD117和CD34在急性白血病中的表达及意义。方法:采用三色流式细胞术,CD45/SSC双参数散点图设门方法,对34例急性白血病患者[急性淋巴细胞白血病(ALL)9例和急性髓细胞白血病(AML)25例]骨髓白血病细胞的免疫分型及CD117和CD34的表达进行检测。结果:CD117在ALL和AML中表达率分别为0和76.0%(19/25),差异有统计学意义(P=0.000)。CD34在ALL和AML中表达率分别为77.8%(7/9)和60.0%(15/25),差异无统计学意义(P=0.815)。在AML-M3亚型中,CD34表达率为16.7%(1/6),显著低于非M3亚型73.7%(14/19)(P=0.007)。结论:CD117检测有助于ALL与AML的鉴别诊断,CD34检测有助于区分AML-M3亚型和非M3亚型。     

15例急性混合细胞白血病的临床与实验研究

目的：研究急性混合细胞白血病(MAL)的临床特征、实验室指标、治疗及预后。方法：对15例MAL骨髓标本分别进行光镜细胞形态学及相关细胞化学染色观察，以确定其FAB类型，应用流式细胞仪做免疫分型检测一系列相关单抗，同时采用G显带技术进行核型分析；应用PCR基因扩增方法检测TCRδ／lgH CDRⅢ2基因重排；运用急性粒细胞白血病(AML)、急性淋巴细胞白血病(ALL)或兼顾二者的治疗方案。结果：MAL患者的临床表现与AML及ALL患者的差异无显著性意义，形态学上MAL表现为AML的多为M2a、M1、M4，表现为ALL的多为ALL-L2；免疫分型显示MAL患者中以三系共表达者多见。另外，CDll7、HLA-DR在MAL中呈高表达。提示MAL的白血病细胞可能起源于早期造血细胞；核型分析可见异常染色体出现，但无特异性；基因重排显示部分患者TCRδ／lgH CDRⅢ2为阳性；患者对治疗反应差，生存期相对较短。结论：MAL具有独特的临床、实验室检查指标及预后。     

格列卫治疗Ph+急性白血病2例报告

费城染色体 (Ph染色体 )虽为慢性粒细胞白血病 (CML)的标志性染色体 ,但也见于 5 %的儿童急性淋巴细胞白血病 (ALL)、1 5 %～ 30 %的成人ALL以及 2 %的急性粒细胞白血病 (AML) 〔1〕。格列卫为针对Ph染色体的分子靶向药物。我院最近采用格列卫治疗Ph+急性白血病 (AL)住院患者 2例 ,现报告如下。1 　 病例报告例 1　男 ,2 9岁。诊断为急性混合细胞白血病。检查骨髓象示原始细胞占 97%。外周血血常规示原始细胞占 74%。采用三色免疫荧光流式细胞术进行骨髓细胞免疫分型 ;以CD45设门 ,病态细胞占 99% ,其中CD1 0 96% ,CD1 999% ,CD33…     

120例白血病细胞免疫表型分析与形态学分析的比较

目的 比较细胞免疫表型分析和形态学分析在白血病诊断和分类中的意义。方法 采用BD公司急性白血病表型分析试剂盒,以流式细胞术对120例白血病患者细胞免疫表型进行分析,并与同期形态学分析结果比较。结果 白血病患者120例,形态学分析正确归类者80例,符合率为66．7％,免疫表型分析可准确对白血病进行诊断和分类。结论 形态学分析和免疫表型分析结合,是白血病诊断和分类的较佳途径。     

Simultaneous analysis of cell surface antigens and DNA content by flow cytometry

Summary Cells obtained from the blood or bone marrow of patients with haematological malignancies can both be stained with fluorescent labelled monoclonal antibodies to determine an immunophenotype and permeabilized with 30% ethanol then stained with propidium iodide for simultaneous DNA analysis. In the technique described here, no evidence of selective depletion of the malignant cell population was demonstrated and decreases in the mean fluorescence intensity of the surface markers were insufficient to affect the sensitivity of flow cytometric analysis. The combined method is simple and robust enough to allow incorporation of DNA analysis into routine immunophenotyping of leukaemia and lymphoma cells.     

A study for evaluation of different diagnostic approaches in acute leukemia in Egypt

Cytomorphology, cytochemistry, immunophenotyping, in addition to cytogenetic and molecular analyses have specific roles in the diagnosis and management of acute leukemias. This work was designed as a comparative study of different available methods for diagnosis of acute leukemia. The study comprised 47 cases with acute leukemia (21 cases with ALL and 26 cases with AML). Peripheral blood and bone marrow samples were subjected to through morphological examination of Leishman-stained smears, cytochemical analysis, immunophenotyping, conventional cytogenetic banding analysis, fluorescence in situ hybridization (FISH) for selected cases, and RT-PCR for detection of BCR-ABL rearrangement. The results of the study revealed that careful examination of Romanowsky-stained peripheral blood and BM films is fundamental in the diagnosis of acute leukemias, and when considered together with clinical and hematological features, indicates which of the more specialized techniques are most likely to be useful. The major role of cytochemistry was in the diagnosis of AML, while the major role of immunophenotyping was in the diagnosis of acute leukemia, which is not obviously myeloid. Apart from identification of chromosomal abnormalities unique to specific subtypes of leukemia, cytogenetic analysis had a salient impact on anticipating the prognosis and treatment outcome in acute leukemias. We could conclude that the techniques used in this study are considered complementary rather than alternatives and that stepwise employment of strategies is more cost effective than doing all the tests simultaneously.     

Acute leukemia subclassification: a marker protein expression perspective

Improved leukemia classification and tailoring of therapy have greatly improved patient outcome particularly for children with acute leukemia (AL). Using immunophenotyping, molecular genetics and cytogenetics the low hanging fruits of biomedical research have been successfully incorporated in routine diagnosis of leukemia subclasses. Future improvements in the classification and understanding of leukemia biology will very likely be more slow and laborious. Recently, gene expression profiling has provided a framework for the global molecular analysis of hematological cancers, and high throughput proteomic analysis of leukemia samples is on the way. Here we consider classification of acute leukemia samples by flow cytometry using the marker proteins of immunophenotyping as a component of the proteome. Marker protein expressions are converted into quantitative expression values and subjected to computational analysis. Quantitative multivariate analysis from panels of marker proteins has demonstrated that marker protein expression profiles can distinguish MLLre from non-MLLre ALL cases and also allow to specifically distinguish MLL/AF4 cases. Potentially, these quantitative expression analyses can be used in clinical diagnosis. Immunophenotypic data collection using flow cytometry is a fast and relatively easily accessible technology that has already been implemented in most centers for leukemia diagnosis and the translation into quantitative expression data sets is a matter of flow cytometer settings and output calibration. However, before application in clinical diagnostics can occur it is crucial that quantitative immunophenotypic data set analysis is validated in independent experiments and in large data sets.     

Quantitative identification of blood cell markers in human hematopoietic malignancies with diagnostic and prognostic significance

Fluorescence intensity (FI) is the basis for classifying phenotypes by fluorescence-label flow cytometry. FI is of a relative value, but with calibration it can be expressed in stoichiometric units called molecules of equivalent soluble fluorochrome (MESF) that reflect the concentrations of the fluorescent conjugates and the receptors they stain. Flow cytometry allows in addition to the determination of positive cells, to establish even the intensity of fluorescent staining, that can be converted into antigen density. The concept of antigen density appears to improve the efficiency of immune techniques in the monitoring of hematopoietic malignancies. Quantitative immunophenotyping is thus suitable for the diagnosis of malignancy, contributes to prognosis and could provide new relevant pathophysiological informations. Quantitative analysis of some markers of leukemic cells could be a good model for the study of antigen modulation caused by chemotherapeutic agens. Standardized reagents and techniques for performing quantitative FI measurements on cytometers are just now emerging into practical use. This latter feature should see the expanding application in both, the basic science and medical applications, as the development of therapeutics increasingly targets specific cell receptors. FI data constitute a very important component of the analysis of cells in hematopoietic malignancy; standardized approach to instrument quality control, interlaboratory comparability of FI measuremant and quality assurance is required. Key words: fluorescence intensity (FI), molecules of equivalent soluble fluorochrome (MESF), quality control, quantitative flow cytometry, leukemia/lymphoma.     

Previously undescribed form of B-cell chronic lymphoid leukemia with IgA expression/secretion and lytic bone lesions

A B-cell chronic lymphoid leukemia (B-CLL) associated with IgA expression and secretion is uncommon and has never been described in association with osteolytic bone lesions. We report such a case, defined by cytomorphology and flow cytometric immunophenotyping (FCI). Additional cases may be recognized with the aid of FCI, in order to define the natural history of and best form of therapy for this rare disorder. Am. J. Hematol. 55:208–211, 1997. © 1997 Wiley-Liss, Inc.     

急性白血病免疫分型中系列相关抗体的敏感性和特异性研究

目的：确定急性白血病免疫分型中系列相关抗体的敏感性和特异性。方法：应用St Jude研究医院的急性白血病免疫分型方案对63例急性白血病患者进行了流式细胞术分型。结果：B系列急性淋巴细胞白血病(B-ALL)敏感性最高的单克隆抗体(抗体)是CD19[100％(22／22)]，而特异性最高的是CD79a[96．7％(29／30)]。CD79a敏感性(95．0％)和特异性均很高，在所有B细胞系列相关抗体中确定系列来源的准确性最高[96％(48／50)]。在T-ALL，敏感性最高的抗体是CD7[100％(14／14)]和CyCD3[100％(12／12)]，特异性最高的是CD5[(100％(45／45)]和CyCD3[97．7％(42／43)]；CD7、CyCD3和CD5在BALL均无假阳性；在AML的假阳性分别是[12％(3／25)]、[4．3％(1／23)]和0／24；但CD5的敏感性仅E85．7％(12／14)]；CD3的敏感性[61．5％(8／13)]和特异性[88．6％(39／44)]均明显低于上述抗体。综合来看，CyCD3敏感性和特异性均很高，在所有T细胞系列相关抗体中确定系列来源的准确性最高[98．2％(54／55)]。在AML，MPO的敏感性和特异性最高[100％(23／23)]和[92．9％(26／28)]，而CD13和CD33的敏感性仅在61．5％和76．9％，特异性仅为68．8％和75．0％。结论：St Jude急性白血病免疫分型方案的高敏感性和高特异性抗体在本组B-ALL和T-ALL病例得以重复；但在AML仅有MPO表现出高敏感性和高特异性；CD13和CD33未能得到高敏感性。     

Impact of CD133 (AC133) and CD90 expression analysis for acute leukemia immunophenotyping

BACKGROUND AND OBJECTIVES: AC133 is a novel monoclonal antibody (Moab) reacting with a population of immature/primitive or granulo-monocytic committed CD34+ve cells. Up to now, only few studies with small numbers of cases have examined AC133 (recently designated CD133) expression in acute leukemia. To determine the value of this Moab for acute leukemia immunophenotyping, we investigated a large series of leukemic cell samples for their reactivity with Moab AC133. DESIGN AND METHODS. A total of 298 cell samples from patients with de novo acute myeloid leukemia (AML) (n=142), acute lymphoblastic leukemia (ALL) (n=119), CD34+ve biphenotypic acute leukemia (n=13), and CD34+ve CML blast crisis (=BC; 21 myeloid BC/3 lymphoid BC) were investigated by flow cytometry for Moab AC133 reactivity.CD133 expression was compared with CD90(Thy-1) expression, another CD34-associated antigen. RESULTS: Fifteen (5%) samples expressed CD90, whereas 114 (38%) samples were positive for Moab AC133 (20% cut-off level). No significant differences in CD133 and CD90 expression levels between AML and ALL were observed. In AML, but not ALL, CD133 was more often expressed in CD34+ve cases than in CD34-ve ones (p<0.00001). However, CD133 expression was not restricted to CD34+ve leukemic cells in individual cell samples. All 8 pro-B-ALL cell samples with 11q23-anomalies and MLL (mixed lineage leukemia) gene translocations were positive for CD133, whereas only 2 of 9 pro-B-ALL without MLL gene translocations expressed CD133 (p<0.002). In contrast, none of the 5 AML cell samples with a t(9;11) and MLL gene translocation reacted with Moab AC133. CD34+ve CML cells in myeloid BC were less often positive for CD133 than CD34+ve de novo AML cells (p<0.0001). INTERPRETATION AND CONCLUSIONS: CD133 and CD90 expression analysis is not helpful for lineage determination in acute leukemia immunophenotyping. However, MoabAC133 may be an informative marker for the detection and further characterization of immature AML cells, as well as pro-B-ALL cells with MLL gene translocations, by flow cytometry.     

A new model of pre-B acute lymphoblastic leukemia chemically induced in rats

OBJECTIVE: Although B acute lymphoblastic leukemia (B-ALL) is the most common leukemia among children, no chemically inducible model of this leukemia has yet been described in vivo. METHODS: Leukemia was chemically induced in male WKAH/Hkm rats by a nitrosourea derivative, N-butylnitrosourea (BNU), an alkylating agent, administered orally 5 days a week for 24 weeks. Development of leukemia was monitored by clinical observation, follow-up of blood parameters, and appearance of blast cells in peripheral blood samples. The phenotype of the leukemia was determined by cytological examination, cytochemical reactions, and by immunophenotyping of bone marrow cells using various markers. The feasibility of leukemia transplantation was investigated. Clonality and karyotype analyses were also performed. RESULTS: We observed the appearance of acute leukemia in 60% of the rats treated with BNU. Of these, 65% developed pre-B-ALL, which was serially transplantable to healthy WKAH/Hkm male rats. Karyotype analysis did not reveal clonal abnormalities. Clonality determined by immunoglobulin gene rearrangement sequencing disclosed that the pre-B-ALL were mostly oligoclonal. CONCLUSION: This new in vivo model of inducible pre-B-ALL might be useful for investigating the effects of co-initiating or promoting agents suspected to be involved in leukemia development, and for disclosing new molecular events leading to leukemogenic processes.