应用一小组抗体对常规切片的分化不良大细胞肿瘤分类——附有随访的免疫组织化学研究

在常规石蜡切片,部分分化不良大细胞肿瘤难以分类。为估计预后和选择恰当的治疗,区分大细胞淋巴瘤、癌、恶性黑色素瘤或肉瘤很重要。虽有报告应用免疫组化法对此类肿瘤进行区别,但尚无用抗体小组直接区分的报告。本文报告用5种抗体构成的小组对29例分化不良大细胞肿瘤分类研究,而这些肿瘤即使有经验的病理学家用常规组织学和组化法,也不能分类。抗体由5种分别为抗角蛋白、波形纤维、人乳汁脂球膜抗原MAM-6、黑色素相关抗原和白血细胞共同抗原的抗体构成,能使用常规固定、石蜡包埋的组织。应用本抗体小组的结果为95％的病例明确诊断;其余病例则得到有价值的信息,可通过辅加标记和电镜观察肯定诊断。随访和治疗结果,与抗体小组的诊断相符。     

恶性淋巴瘤切片诊断新方法

在常规肿瘤病理检验中，有时候由于组织固定不适当，会出现淋巴瘤切片染色不够清晰的问题，影响了恶性淋巴瘤的诊断工作。我们参考国外献：采用B5固定液重新固定组织切片的方法，改善了淋巴瘤组织切片的微细结构，使不易诊断的淋巴瘤得到了诊断。现将该法介绍如下。     

超声波快速制片法的应用

超声波快速制片法的应用蔡俊杰丁彦青张盛在临床工作中，常常会遇到需要快速明确病理诊断的病人，以往是行冰冻切片或人工快速石蜡切片而作出病理诊断，但冰冻切片不宜长期保存，诊断的可靠性不如石蜡切片，人工快速石蜡切片费工费时，工作量大时不易做到。为使病人能及时...     

胃MALT淋巴瘤免疫组化及病理形态定量研究

目的：探讨胃粘膜相关淋巴组织（ＭＡＬＴ）诊断与分类方法。方法：对１４例ＭＡＬＴ淋巴瘤进行免疫组化及病理形态定量研究。标本采用常规石蜡切片、ＨＥ染色及ＡＢＣ法免组化染色,并应用ＨＰＬＡＳ－１０００彩色图像分析系统对肿瘤细胞核的形态进行定量测定。结果：１４例胃ＭＡＬＴ淋巴瘤中ＣＣＬ细胞性９例。小无裂、大无裂、Ｔ免疫母细胞性各１例,ＣＣＬ合并大细胞性淋巴瘤２例。形态定量测定结果表明胃ＭＡＬＴ淋巴瘤与对照     

术中冰冻切片对卵巢肿瘤诊断的价值

目的：对２４８例卵巢肿瘤的临床诊断及术中冰冻切片（ＦＳ）诊断与石蜡切片诊断进行对比研究。方法：石蜡切片诊断良性肿瘤１１８例，交界性肿瘤８例，恶性肿瘤７２例。临床诊断和石蜡切片符合率为７５．８％（１８８／２４８），良性肿瘤为９６．６％（１１４／１１８），交界性肿瘤为４８．３％（４７／５８），恶性肿瘤为６５．３％（４７／７２）。结果：ＦＳ诊断与石蜡切片的符合率为９０．３％（２２４／２４８），良性肿瘤为     

在B-5和福尔马林固定的石蜡切片中鉴别滤泡性淋巴瘤和滤泡反应性增生

对石蜡包埋常规切片组织学区别滤泡性淋巴瘤和滤泡性反应性增生,常常是相当困难的,甚至单克隆免疫表型也难以将二者容易地鉴别。为寻找一个单克隆抗体能在石蜡组织切片中对淋巴样细胞起反应,并用于鉴别证实细胞的增殖性质,作者用下列抗体检测了45例滤泡淋巴瘤和30例滤泡反应性增生。其抗体为:L_(26),B_2,MT_1,MT_2和UCHL-1,所有切     

免疫组化在霍奇金淋巴瘤诊断中的应用

近年的研究发现经典型霍奇金淋巴瘤(CHL)、结节性淋巴细胞为主型的霍奇金淋巴瘤(NLPHL)以及富于T细胞B细胞淋巴瘤(TCRBCL)间存在一个诊断灰区(gray zone)[1～5]。为了进一步了解石蜡切片免疫组化在霍奇金淋巴瘤(HL)诊断方面的作用，我们回顾性地分析1995～1999年间22例HL的诊断，现报告如下。 1 材料和方法 1.1 材料 22例均是我院病理活检病例，采用常规形态学诊断。 1.2 方法 免疫组化采用SP法，在4μm石蜡切片上进行。微波抗原修复，应用Dako公司提供的抗CD20(L26)、CD30(Ber-H2)、CD15(C3D-1)CD21(1F8)、CD45(PD7/26)、Vimentin(V9)、CD45RO(UCHL-1)和CD57(Leu7)抗体。 1.3 分类 按照REAL和WHO分类标准[4,5],进行重新评价。     

印片加微波快速切片在卵巢肿瘤术中诊断的应用

目的：探讨印片加微波快速石蜡切片在卵巢肿瘤术中诊断的应用及准确性问题。方法：收集82例卵巢肿瘤术中印片加微波快速石蜡切片诊断与常规石蜡切片诊断资料,将卵巢肿瘤分为瘤样病变、良性肿瘤、交界性肿瘤及恶性肿瘤四类进行对照研究。结果：82例中确诊79例，确诊率96．34％（其中完全符合61例，基本符合18例）；未能确诊3例，占3．66％。对卵巢瘤样病变、良性肿瘤、交界性肿瘤和恶性肿瘤的确诊率分别为100％、97．22％、83．33％。结论：应用印片加微波快速石蜡切片可取两者之长，提高诊断的准确性、减少误诊，不失为术中诊断的一种较好方法。     

某些单(多)克隆抗体在肿瘤鉴别诊断中的应用(97例疑难病例分析)

应用免疫组织化学的方法,藉助于单(多)克隆抗体,在组织切片上确定特定抗原的部位,97例疑难肿瘤病例得到分析,此研究显示,角蛋白和上皮细胞膜抗原在证明未分化癌中颇有用处,恶性淋巴瘤能够应用白细胞共同抗原来确诊,而S100蛋白能够确定神经纤维瘤和恶性黑色素瘤,等等;本研究提示,单(多)克隆抗体应用于肿瘤的鉴别诊断中,具有重要意义。     

骨髓切片和涂片观察在恶性血液病诊断中的意义

 目的 应用骨髓切片和涂片相结合,观察骨髓组织和细胞形态学的变化,对120例初发的血液病患者的骨髓进行切片和涂片的结果分析,以期得到形态学诊断的新途径。方法 收集恶性血液病病例120份。一步法抽吸骨髓,常规方法涂片染色,塑料包埋法制作组织切片;恶性血液病的诊断参照2001年WHO分型标准。结果 骨髓活检与骨髓涂片相结合,在诊断骨髓纤维化、低增生MDS、骨髓转移癌和骨髓坏死,骨髓组织切片的诊断符合率高于穿刺涂片。对于淋巴瘤骨髓侵犯的病例,骨髓组织切片中更容易找到有特征的淋巴瘤细胞;对慢性粒细胞白血病（CML）病例,组织切片较穿刺涂片提前找到诊断急变的证据。结论 骨髓形态学检验,将组织切片与穿刺涂片相结合,互为补充,可以有效地提高恶性血液病的诊断水平。     

Different antigen unmasking techniques lead to significant differences in immunohistochemical staining of Ki-67 (Mib-1) in renal cell carcinomas.

For immunohistochemistry on formalin-fixed, paraffin-embedded specimens adequate pretreatment of the sections to achieve an unmasking of antigenic domains before staining the specimens is necessary. In this study, we compared the effect of microwave and autoclave heating on the expression of the cell proliferation marker Ki-67 (Mib-1) in renal cell carcinomas. A total of 177 formalin-fixed and paraffin-embedded tumor specimens of locally confined renal cell carcinomas with clear cell morphology were investigated. Two different antigen unmasking techniques were used: microwave and autoclave heating. The fraction of Ki-67 positively stained nuclei was determined in APAAP (alkaline phosphatase-antialkaline phosphatase) stained slides. The fraction of Mib-1-labelled cells was significantly higher after autoclave compared to microwave pretreatment (mean value 4.95 vs. 1.43, p<0.001). We recommend the autoclave heating technique for quantitative detection of Ki-67 with the Mib-1 antibody in renal cell carcinomas.     

Significance of wet autoclave pretreatment in immunohistochemistry

Until recently the only way to rescue masked epitopes in routinely processed surgical pathological material was enzymatic digestion. The use of heat for antigen retrieval, first by microwave irradiation, represents an important breakthrough in immunohistochemistry. With the acceptance of microwave oven pretreatment, various modified techniques and alternative heating methods have also been proposed. Wet autoclave pretreatment for tissue proteolysis is a highly reliable alternative to the microwave antigen retrieval technique. It provides uniform heating of the slides, hence an even enhancement of staining intensity in a variety of formalin-sensitive antigens, and it also offers consistent interlaboratory results. The method has been introduced in routine diagnostic immunohistochemistry for the detection of estrogen-and progesterone receptors, L26-, Ki-67- and bcl-2 antigens and variable types of cytokeratins (1/5/10/11, 8, 13, 19). Experimentally, wet autoclaving can be used very successfully for the immunophenotyping of p53 and mdm2 expression, for the detection of adhesion molecules (CD44, integrins) and some anti-inflammatory molecules (annexins), among others. It has produced a substantial improvement in the visualisation of silver-stained nucleolar organizer regionsassociated proteins (AgNORs) in routine paraffin sections and along with modified silver staining and standardized AgNOR parameters assessed by image analysis. Wet autoclaving-based AgNOR staining has been proposed by a European multicentric study group as the standardized method for AgNOR analysis in archival material.     

Immunostaining of human glioblastomas with various PCNA antibodies

The aim of the study was to compare various PCNA clones (PC10, 19A2, and 19F4) on frozen and paraffin wax sections of ten human glioblastomas. Standard immunohistochemical methods were used (avidin-biotin-peroxidase technique). Half of the paraffin sections were pretreated in a microwave oven. With few exceptions positive staining was achieved only on paraffin sections after microwave treatment. Staining with the 19A2 antibody revealed few (<1%) positively stained tumor nuclei in only three cases whereas staining with 19F4 was positive in eight cases with a labeling index (LI) in the range of <1-3.5%. All cases showed positive staining with the PC10 antibody with an LI of <1% in three cases and in the range of 2.0-17.2% in the remaining seven. However, most of the PC10 stained sections showed a disturbing unspecific background staining. Thus, in spite of the background staining we conclude that PC10 offered the best immunostaining of the antibodies tested.     

Immunohistochemical demonstration of estrogen receptors on routine paraffin sections of breast carcinomas: a comparison with frozen sections and an enzyme immunoassay.

In this study the monoclonal antibody ER-ICA (HSpy222) to human estrogen receptor (ER) protein and the peroxidase-antiperoxidase method was used to detect the presence of ER in 83 cryostat sections and in 68 paraffin sections pretreated with pronase in a total of 86 primary breast cancers. In 72 out of the 86 studied cases, a comparative evaluation was performed between the semiquantitative ER-ICA method and the quantitative enzyme immunoassay ER-EIA. A good correlation was found between the semiquantitative ER-ICA results in cryostat and paraffin sections (95.38%; p less than 0.01) in a total of 65 compared cases, concerning both the percentage of ER-positive or negative cells and the staining intensity. In addition, the overall appraisal of the lesion as ER-ICA-positive or ER-ICA-negative as well as the ER-ICA staining intensity and the proportion of ER-ICA stained cancer cells, in both cryostat and paraffin sections, correlated significantly with the mean values of fmol ER/mg determined by the enzyme immunoassay ER-EIA. The performance of the ER-ICA method on paraffin sections as used in the present study proved to be a reliable and reproducible immunohistochemical technique.     

Loss of antigenicity in stored sections of breast cancer tissue microarrays.

Immunohistochemical characterization of tumor tissues in epidemiological studies is a promising approach to identify breast cancer subtypes with distinct etiology. The recent development of the tissue microarray (TMA) technique allows for standardized, rapid, and cost-effective immunohistochemical characterization of many cases, which is critical in epidemiological studies. Sectioning paraffin blocks at different times results in loss of material, which can be reduced by preparing many sections each time a block is cut. However, data suggest that staining intensity declines in whole sections prepared from conventional paraffin blocks with storage time, resulting in false-negative results. This problem would be accentuated in TMAs because of the limited tissue representation of each case. To evaluate this concern, we prepared a single TMA block from 125 invasive breast carcinomas collected in a population-based case-control study conducted in Poland and compared estrogen receptor (ER-alpha), progesterone receptor (PR), and human epidermal growth factor receptor 2 (HER2) expression in sections cut and stored for 6 months at room temperature with sections cut from the same TMA block and stained on the same day. Percentage of positive cases for stored versus fresh sections was similar for ER (59.0%) but significantly higher in fresh sections for PR (56.3% versus 64.1%, P = 0.01) and HER2 (45.5% versus 64.4%, P < 0.001). Among cases positive in both stored and fresh sections, the median percentage of immunoreactive cells was significantly reduced and the staining intensity was consistently lower in stored compared with fresh sections. We conclude that loss of immunoreactivity is an important problem in TMAs of breast cancer. Improved methods for sectioning TMAs and storing tissue sections aimed at reducing loss of immunoreactivity are critical for the use of TMAs in epidemiological studies.     

Immunohistochemical detection of estrogen receptors on paraffin sections of normal, hyperplatic and carcinomatous endometrium.

The present study is the first dealing with the demonstration of estrogen receptors (ER) in up to 8-year-old paraffin blocks of endometrial curettage samples routinely fixed in 10% formalin. The Mab ER-ICA was used in a modified peroxidase-antiperoxidase method after pretreatment of paraffin sections with pronase. Eleven cases with proliferative, 11 cases with secretory endometrium, 20 cases with adenocystic, 21 with adenomatous hyperplasia and 27 endometrial adenocarcinomas were tested. The two main parameters, namely the percentage of ER-positive cells and the intensity of the immunostaining, were higher in the proliferative phase followed in a declining sequence by adenocystic hyperplasia, adenomatous hyperplasia, adenocarcinomas and the secretory phase of endometrium. Interestingly, the intensity of the immunostaining showed a positive relationship to the percentage of ER-positive cells (r = 0.93, p less than 0.001). It seems that the immunohistochemical demonstration of ER in paraffin sections of uterine specimens is an easy and reliable method for the mapping of the heterogeneous expression of ER and their comparative study with the well preserved histopathological features even in old archival paraffin-embedded material.     

Rapid immunohistochemical detection of tumor cells in gastric carcinoma

The detection of single tumor cells or tumor cell clusters represents an important issue in intraoperative frozen section analysis. For example, surgical margins may be evaluated in order to minimize the number of additional operations. Furthermore, intraoperative diagnosis of lymph node micrometastasis (LNM) may help to define the area of appropriate lymph node dissection. In addition to haematoxylin and eosin (H&E)-stained sections, immunohistochemical detection of single tumor cells or cell clusters may be helpful in this context. The aim of this study was to evaluate the clinical significance, reliability and sensitivity of intraoperative rapid immunostaining of frozen sections. Therefore, we compared the results of rapid immunohistochemical staining of frozen sections and paraffin sections applying the EnVision and Histofine(R) detection systems. In a prospective immunohistochemical study, paraffin and frozen sections of 20 gastric cancer specimens were analyzed. Paraffin as well as frozen sections were stained immunohistochemically applying the EnVision and Histofine detection systems. As primary antibodies, AE1/AE3 (anti-cytokeratin), EMA (anti-MUC1) and B lymphocyte marker anti-CD20 were applied. The rapid immunostaining procedure was able to be completed within 10-13 min. Rapid immunohistochemical staining of frozen and paraffin sections of the same tumors resulted in comparable immunoreactivity. The rapid EnVision and Histofine procedures allowed immunostaining of frozen sections in less than 13 min. These methods can represent useful additional tools in routine surgical pathology and research, enabling a more accurate frozen section diagnosis compared to staining with H&E alone. Intraoperative rapid immunostaining can be a simple and useful technique to detect LNM.     

Colocalization of estrogen and progesterone receptors with an estrogenregulated heat shock protein in paraffin sections of human breast and endometrial cancer tissue

Summary We have studied by immunocytochemistry and monoclonal antibodies the presence and localization of estrogen receptors, progesterone receptors, and a 24-kD estrogen-regulated heat shock protein in biopsies from breast and endometrial cancer patients. Three different tissue processing protocols were used to colocalize the antigens in the same tissue sections: a) frozen sections, b) formalin fixation with routine paraffin embedding, and c) picric acid-formaldehyde (PAF) fixation with a rapid embedding in paraffin. Frozen sections showed good receptor staining but poor 24-kD protein immunoreactivity, while routine paraffin sections (with or without DNase pretreatment) were inadequate to reveal the nuclear receptor proteins at the same level seen in frozen sections. On the other hand, all three proteins could be detected satisfactorily in PAF-fixed paraffin-embedded tissue. Using this procedure we were able to visualize 24-kD protein and estrogen receptor or progesterone receptor in individual cells in paraffin sections. The study revealed that in all of the estrogen receptor positive breast and endometrial tumor samples, almost 90% of the cells expressing the cytoplasmic 24-kD protein contained estrogen receptor in the cell nucleus. In contrast, 24-kD immunoreactive cells did not express progesterone receptors in almost 40% of the progesterone receptor positive tumor samples.     

切片位置对兔睾丸组织体视学研究的影响

目的比较不同包埋介质内切片位置对成年新西兰大耳白兔睾丸组织体视学研究的影响。方法将成年雄性新西兰大耳白兔麻醉后,随机抽选一侧睾丸经Bouin液固定后从中央及两端（垂直于睾丸中心长轴）切取共三个平行组织薄片。将每个组织薄片平分为二,再随机将其中一块平分得到2个（1/4圆）组织块,分别随机采用羟乙基甲基丙烯酸树脂或石蜡包埋、切片和染色后,在低倍镜（10×）下分别对睾丸组织树脂切片（25μm）和石蜡切片（7μm）不同切片位置进行观察：紧贴白膜选择若干视野（所有视野间无任何重叠）,再向扇形切片中心连续选择3个视野,利用体视学方法估计各视野睾丸组织内生精小管的体积分数。结果石蜡切片较树脂切片估计的生精小管体积分数减少15.9%;树脂切片不同部位估计的生精小管体积分数无差异,而石蜡切片不同部位估计的结果存在差异：距离白膜愈远,体积分数愈小,近切片中心与近白膜相比显著减少了30.1%。结论石蜡包埋能引起睾丸组织不均匀皱缩,因此,当应用石蜡切片进行睾丸体积分数研究时,应考虑不均匀组织皱缩因素。     

Expression of epidermal growth factor receptor (EGF-R) in non-small cell lung cancer. Use of archival tissue and correlation of EGF-R with histology, tumour size, node status and survival

A total of 152 non-small cell lung cancers (NSCLC) were studied retrospectively to determine the relationship between epidermal growth factor receptor (EGF-R) status and the histological type, tumour size, nodal status and prognosis. EGF-R status was assessed on routinely embedded paraffin sections with an antibody to the cytoplasmic domain of the tumour (F4 antibody). EGF was demonstrated in all tumour types and every squamous and large cell carcinoma was positive for the antibody. Most tumours showed heterogeneity of staining. EGF expression was seen statistically more frequently in well differentiated tumours. Patients with 50% or more tumour cells showing positivity tended to have an improved survival but this result failed to reach statistical significance. There was no relationship between the size of the primary tumour or the lymph node status. Other cells, such as mucinous glands, bronchial epithelial cells and macrophages stained positively with the monoclonal antibody. EGF receptor status, with the antibodies presently available, adds little to help in either diagnosis or prognosis. Interpretation of data has to be guarded since the antibody was seen in some normal cells.