合成多肽制备的抗体用于蛋白质免疫组织细胞化学定位研究

对本实验室克隆的小麦春化相关基因VER2编码蛋白的氨基酸序列进行软件分析 ,确定了具有免疫原性的一段 (2 0 4～ 2 1 5位 )含 1 2个氨基酸的短肽 ,用化学法合成了该短肽并将其与牛血清白蛋白 (BSA)相偶联 ,用偶联物作为抗原免疫家兔获得该 1 2肽的抗体 .分别用VER2全蛋白抗体和该 1 2肽抗体对VER2蛋白进行免疫细胞定位比较分析 ,两者所得结果一致 ,即二者在免疫组化检测蛋白质时的灵敏性和特异性方面具有一致的特性 .结果表明 ,化学法合成短肽制备的抗体可用于蛋白质的免疫组织细胞化学分析 ,该方法快捷灵敏 。

免疫组织细胞化学在法医学上的应用(文献综述)

法医学的产生和发展从来就是与其他学科，尤其是生物学及医学科学的发展紧密相关的．随着近年来免疫组织化学(Immunohistocytochomistry．简称IHCC)的迅速发展，它已经开始，并将越来越深入地推动法医学各学科的前进，使法医学在深度和广度上以一种崭新的面貌，更好地发挥其为刑事侦察和司法审判服务、以及推动其他有关医学科学发展的作用．

大肠癌组织P21蛋白的表达研究

研究 P2 1蛋白的表达与大肠癌发生的关系。应用免疫组化 SABC法分别检测大肠癌组织及其癌瘤手术切除的切缘组织 P2 1蛋白的表达。P2 1蛋白在大肠癌组织中的阳性率为 6 5 .0 % ,在切缘组织中的阳性率为 15 .0 % ,大肠癌组织与切缘组织阳性率比较有极显著性差异 ,P<0 .0 1。结果表明 P2 1蛋白的表达与大肠癌的组织学类型、 Dukes分期、分化程度无关 ,P>0 .0 5。提示 P2 1蛋白在大肠癌的发生中发挥重要作用。

VEGF在前列腺腺癌和前列腺增生组织中的表达及其意义

目的 探讨血管内皮生长因子在前列腺癌和前列腺增生发生与发展过程中的潜在生物学作用及其与肿瘤细胞分化程度的关系。方法 采用免疫组织细胞化学SABC技术 ,对 31例前列腺癌和 17例前列腺增生组织中血管内皮生长因子的表达与分布状况及其与肿瘤细胞分级的关系进行了检测与分析。结果 血管内皮生长因子在前列腺癌和前列腺增生组织中均有表达 ;在前列腺上皮细胞胞浆、基质细胞和间质中呈弥散分布 ,血管上皮细胞及其周围基质细胞呈强阳性表达。半定量分析结果显示 ,前列腺癌组织中表达强度高于前列腺增生组织 ;未发现其表达强度与前列腺癌细胞分级 (或分化程度 )有统计学关联。结论 该研究提示 ,血管内皮生长因子在前列腺癌和前列腺增生组织中分布广泛 。

IGF-1R在前列腺腺癌和前列腺增生组织中的分布和表达

目的 探讨IGF - 1R在前列腺腺癌和前列腺增生组织中的分布及其细胞生物学意义。方法 采用免疫组织化学技术ABC方法 ,对 2 3例前列腺腺癌和 15例前列腺增生组织中IGF - 1R的表达情况进行了检测分析。结果 前列腺腺癌组织和前列腺增生组织中IGF - 1R表达均呈阳性 ,IGF - 1R主要分布于前列腺上皮细胞膜 ,基质细胞中亦有散在、少量的分布 ;半定量分析结果显示 ,前列腺增生组织中IGF -1R表达 (或染色 )强度高于前列腺癌组织 ;ICF - 1R表达强度与前腺癌细胞分级呈负相关趋势 ,肿瘤细胞分级越高 (或分化越差 )IGF - 1R表达越弱。结论 研究提示 ,IGF - 1R (或IGFs系统 )在前列腺上皮细胞转化和肿瘤细胞增殖分化过程中具有一定作用。

NAP-2和PF-4在烧伤后大鼠骨髓巨核细胞表达变化情况

目的观察大鼠30%Ⅲ度烧伤后趋化因子NAP-2和PF-4在大鼠骨髓巨核细胞的表达变化情况。方法采用免疫组织细胞化学的方法观察烧伤后24h和15d NAP-2和PF-4表达情况。结果大鼠烧伤后24h骨髓巨核细胞特异分泌的趋化因子NAP-2和PF-4明显低于正常组,烧伤后15d巨核细胞表达NAP-2和PF-4较烧伤后24h恢复,但仍低于正常对照组。结论严重烧伤可以导致大鼠骨髓巨核细胞分泌特异趋化因子NAP-2和PF-4的能力减退。

黄芪注射液抗心肌细胞再灌注损伤的作用及机制

该研究采用离体兔心缺血再灌注模型和培养心肌细胞缺氧复氧模型，从器官和细胞两个水平，运用免疫组织细胞化学、流式细胞仪、免疫印记、RT-PCR，生化学检测等多种方法，较系统地研究了黄芪注射液抗再灌注损伤的作用和机制。特别是该论文从细胞信号转导角度研究了黄芪药物作用的分子药理机制．发现该药物具有调节抗再灌注损伤的MAPK细胞信号通路的作用．而这种作用很可能是其心肌保护效应的机制之一。并且特异性抑制剂并不能减弱黄芪作用，说明黄芪可能是通过多种途径发挥作用。

庆大霉素损伤后豚鼠前庭上皮自发修复的分子机制研究

目的观察庆大霉素损伤后豚鼠前庭上皮自发修复期细胞增殖和胞外信号调控蛋白激酶1/2(extracellularsignalreg-ulatedkinase1/2,ERK1/2)的表达和分布特点,探讨其生物学特性和意义。方法采用免疫组织细胞化学方法,以ERK1/2抗体为标记物,检测庆大霉素损伤后豚鼠前庭上皮修复期ERK1/2的表达变化和分布特点;采用Brdu体内标记和抗Brdu抗体免疫组化染色,检测庆大霉素损伤后豚鼠前庭上皮修复期细胞增殖情况。结果ERK1/2在正常对照组前庭上皮细胞中有低水平表达。庆大霉素损伤后,其表达渐增。在损伤后1d时ERK1/2表达明显增多,7d时表达最强,其后逐渐减少。在正常对照组前庭上皮中无Brdu阳性细胞。各实验组均观察到Brdu阳性细胞,7d时最多。结论庆大霉素损伤后,豚鼠前庭上皮ERK1/2表达增强,其时程变化与细胞增殖活动一致。ERK1/2在豚鼠前庭上皮损伤后的自发修复中可能起重要信号转导作用。

血管内皮生长因子及其受体在鼻息肉中的表达

目的 :检测血管内皮生长因子及其受体在鼻息肉中的表达。方法 :将正常鼻粘膜和鼻息肉用 10 %福尔马林固定 ,石蜡包埋 ,切片。用免疫组织化学方法检测血管内皮生长因子及其受体在正常鼻粘膜和鼻息肉中的表达。结果 :血管内皮生长因子及其受体在正常鼻粘膜呈弱阳性 ,在鼻息肉呈强阳性表达。结论 :血管内皮生长因子在鼻息肉的形成中起重要作用。

Negative correlation of Nogo-A with the malignancy of oligodendroglial tumor

Objective Nogo-A is an axon regeneration inhibitor, and its function in central nervous system （CNS） is still unknown. The present study is to explore the relationship between the expression of Nogo-A and the malignancy of oligodendroglial tumors in patients. Methods Tumor tissue samples with different malignancy grade were obtained from the hospitals. The samples used for detection had been diagnosed as oligodendroglial tumors （oligodendroglioma or anaplastic oligodendroglioma）. The expression of Nogo-A was detected by irmnunohistochemistry and western-blot analysis. The correlation test between the Nogo-A expression and the morphological changes （the percentages of atypical cells and mitotic cells in the tumors） related to the malignancy of tumor tissues was performed. Results There was significant negative correlation between the Nogo-A expression and the morphological change of tumor tissues according to immunohistochemistry. Western-blot analysis also indicated that the gray value of Nogo-A protein band in the oligo- dendroglioma group was significantly higher than that in the anaplastic oligodendroglioma group. Conclusion Nogo-A expression was negatively correlated with the malignancy grade of oligodendroglial tumors.

Cell Cycle Kinetic Analysis in the Cortical Regions of the Lentil Primary Root During Germination

Cell cycle kinetic activity in the cortical cells of the lentil (Lens culinaris Me-die. cv. Verte du Puy) primary root during germination was examined both temporally and spatially. Immunohistochemical and cytological evidence indicated that DNA replication and cell division started in the cortical cells of tire lentil primary root after around 13 and 17 h of imbibition, respectively. The first cells in DNA synthesis and the First mitotic figures all appeared in the cortical cells about I mm front the root-cap junction, but these divided cells had synthesized their DNA during the maturity of seed instead of during germination. The kinetic pattern of activity of the first cell cycle showed that these cells were not activated synchronously, but re-entered the cell cycle in turn depending on their places in the root tip, However, the adjacent cells partially synchronously proceeded their cell cycle.

The Expression of RECK mRNA and Protein in Esophageal Squamous Cell Carcinoma and Its Clinical Significance

OBJECTIVE To investigate the expression of RECK mRNA and protein in esophageal squamous cell carcinoma （ESCC） and to examine its relationship with the clinicopathologic features. METHODS The expression of RECK mRNA and protein in 62 cases of ESCC, 31 of paraneoplastic atypical hyperplasia （PAH） and 62 normal esophageal mucous membrane specimens was examined, using RT-PCR and immunohistochemistry. RESULTS During canceration of the ESCC, the mRNA of RECK increased sequentially from ESCC tissue to PAH and normal mucous membranes. Values were 1.052±0.078, 1.274±0.235 and 1.306±0.121, respectively, with a significant difference among different groups （F=49.936, P〈0.05）. There was a statistically significant difference in the relative amount of the RECK mRNA among the ESCC tissues at various levels of differentiation, depth of infiltration, and different types of lymph node metastasis （F=5.081, F=26.084, U=24.011, P〈0.05）. In the ESCC tissue and PAH, the positive rates of RECK protein expressions were lower compared to the normal mucosa tissue, i.e. 59.7% （37/62）, 71.0% （22/31） and 85.5% （53/62）, respectively. There was a significant difference among the inter-group comparisons （Х^2=10.331, P〈0.01）. In ESCC, there was a close correlation between the RECK protein expression and the degree of cancer differentiation, and the depth of invasion and the types of ESCC lymph node metastasis （P〈 0.05）. CONCLUSION The decrease in expression of both RECK mRNA and protein in ESCC suggest that these low expressions may relate to ESCC development. Examination of RECK mRNA and protein expression may develop into one of the molecular indices for early ESCC diagnosis and prognosis.

Immunohistochemical assessment of angiogenesis in hepatocellular carcinoma and surrounding cirrhotic liver tissues

AIM: To investigate whether vascular endothelial growth factor (VEGF) was over-expressed in hepatocellular carcinoma (HCC) or in surrounding cirrhotic liver tissues.METHODS: Immunohistochemistry was performed to investigate the expression of VEGF proteins in HCC tissues from 105 consecutive patients undergoing curative resection for HCC. The immunostaining results and related clinicopathologic materials were analyzed with statistical methods. Kaplan-Meier method was used to calculate survival curves, and Log-rank test was performed to compare differences in survival rates of the patients with positive HCC staining and negative VEGF.RESULTS: VEGF-positive expression was found in 72 of105 HCC patients (68.6%). Capsular infiltration (P= 0.005),vascular invasion (P = 0.035) and intrahepatic metastasis(P=0.008) were observed more frequently in patients with VEGF-positive expression than in those with VEGFnegative expression. Kaplan-Meier curves showed that VEGF-positive expression was associated with a shorter overall survival (P = 0.014). VEGF-positive expression was found in 47 of tissues 68 HCC (69.1%), and VEGF-positive expression was found in 54 of 68 surrounding cirrhotic liver tissues (79.4%). VEGF-positive expression was significantly higher in surrounding cirrhotic liver tissues than in HCC (P= 0.017).CONCLUSION: VEGF may play an important role in the angiogenesis and prognosis of HCC, as well as in the angiogenesis of liver cirrhosis.

Identification of zRAP55,a gene preponderantly expressed in StagesⅠandⅡ oocytes of zebrafish

In an in silico search for gonand specific expressed genes, we have identified zRAP55 which is enriched in the ovary of zebrafish . zRAP55 encodes a protein of 382 amino acids with a highly conserved Lsm domain. zRAP55 protein shares more than 56% identities with that of other vertebrate species. RT-PCR results show that it is predominantly expressed in the ovary. In situ hybridization and immunohistochemistry studies reveal that zRAP55 is ubiquitously dispersed throughout the cytoplasm of stages Ⅰ and Ⅱ oocytes, whereas no expression is observed in stages Ⅲ and Ⅳ oocytes. As an RNA associated protein, zRAP55 might function in the control of protein translation at the early stages of oogenesis in zebrafish.

Hepatic non-parenchymal cells and extracellular matrix participate in oval cell-mediated liver regeneration

AIM： To elucidate the interaction between non- parenchymal cells, extracellular matrix and oval cells during the restituting process of liver injury induced by partial hepatectomy （PH）. METHODS： We examined the localization of oval cells, non-parenchymal cells, and the extracellular matrix components using immunohistochemical and double immunofluorescent analysis during the proliferation and differentiation of oval cells in N-2- acetylaminofluorene （2-AAF）/PH rat model. RESULTS： By day 2 after PH, small oval cells began to proliferate around the portal area. Most of stellate cells and laminin were present along the hepatic sinusoids in the periportal area. Kupffer cells and fibronectin markedly increased in the whole hepatic Iobule. From day 4 to 9, oval cells spread further into hepatic parenchyma, closely associated with stellate cells, fibronectin and laminin. Kupffer cells admixed with oval cells by day 6 and then decreased in the periportal zone. From day 12 to 15, most of hepatic stellate cells （HSCs）, laminin and fibronectin located around the small hepatocyte nodus, and minority of them appeared in the nodus. Kupffer cells were mainly limited in the pericentral sinusoids. After day 18, the normal liver Iobule structures began to recover.CONCLUSION： Local hepatic microenvironment may participate in the oval cell-mediated liver regeneration through the cell-cell and cell-matrix interactions.

Persistent CXCR4 expression after preoperative chemoradiotherapy predicts early recurrence and poor prognosis in esophageal cancer

AIM: To study the effect of CXC chemokine receptor-4 (CXCR4) expression on disease progression and prognosis in esophageal cancer. METHODS: CXCR4 expression was evaluated in 37 patients with histologically confirmed esophageal squamous carcinomas (ESCC) undergoing preoperative chemoradiotherapy (CRT) by immunohistochemical staining. RESULTS: Eleven out of 37 ESCC patients showed a pathological complete response (CR) after CRT. CXCR4 protein expression was observed in cell cytoplasms of 13 tumors, and null expression was seen in 13 tumors. Distant recurrence was significantly more common in patients with positive CXCR4 expression (P = 0.0318). After a median follow-up time of 31.6 mo, 19 patients progressed (12 of 19 expressed positive CXCR4) and 11 died (10 of 11 expressed positive CXCR4). Overall survival was significantly correlated with lymph node metastasis (952.1 ± 53.8 d in negative group vs 475.1 ± 56.2 d in positive group, P = 0.023), distant metastasis (874.0 ± 60.4 d in negative group vs 434.9 ± 75.2 d in positive group, P = 0.014) and CRT (811.5 ± 51.2 d in responder group vs 459.6 ± 94.0 d in non-responder group, P = 0.00038) and further with an absence ofCXCR4 expression or no residual tumor (959.8 ± 51.0 d in null expression or no tumor group vs 412.0 ± 57.1 d in positive expression group, P = 0.0001). CONCLUSION: Persistent positive CXCR4 expression is implicated in tumor aggressiveness and poor prognosis in ESCC after CRT, and preoperative CRT may improve the prognosis of ESCC via CXCL12-CXCR4 signaling pathway.

Nuclear β-catenin expression as a prognostic factor in advanced colorectal carcinoma

AIM： To investigate the changing pattern of β-catenin expression and its prognostic value in advanced colorectal cancer （CRC）. METHODS： Archival tumor samples were analyzed for β-catenin using immunohistochemistry （IHC） in 95 patients with advanced CRC. RESULTS： Membranous β-catenin expression was found in the normal colorectal epithelium. Almost 100% of CRC cases showed membranous and cytoplasmic expression, and 55 （58%） cases showed nuclear expression. In univariate （Kaplan-Meier） survival analysis, only the nuclear index （NI） was a significant predictor of disease free survival （DFS） （P = 0.023; n = 35）, with a NI above the median associated with longer DFS （34.2 too） than those with a NI below the median （15.5 too） （P = 0.045, ANOVA）. The other indices were not significant predictors of DFS, and none of the three tested indices （for membranous, cytoplasmic, or nuclear expression） predicted diseasespecific survival （DSS）. However, when dichotomized as positive or negative nuclear expression, the former was a significant predictor of more favorable DFS （P = 0.041） and DSS （P = 0.046）. CONCLUSION： Nuclear β-catenin expression provides additional information in predicting patient outcome in advanced CRC.

Reactive lymphoid hyperplasia of the liver:A case report and review of literature

A case of a 53-year-old female patient with reactive lymphoid hyperplasia （RLH）, clinically designated as pseudolymphoma of the liver is described in this article. The patient was admitted to our hospital for further evaluation of hepatic tumors incidentally discovered at another hospital. Various diagnostic methods, including ultrasonography （US）, computerized tomography （CT）, magnetic resonance imaging （MRI） and hepatic angiography displayed three small lesions in the liver with outstanding findings consistent with hepatocellular carcinoma （HCC）. Surgical resection was performed and the three lesions were microscopically diagnosed as RLH of the liver. The lesions comprised a massive infiltration of lymphoid cells with follicles and hyalinized inter- follicular spaces. Immunohistochemical examination revealed that infiltrating lymphocytes had no prominent nuclear atypia and polyclonality. RLH of the liver is a very rare condition and only twelve cases have been reported in the English literature. Majority of the reported cases were middle-aged women and about half of them had some immunologic abnormalities such as autoimmune thyroiditis, Sjogren＇s syndrome, primary immunodeficiency, primary biliary cirrhosis. Since they are olden clinically misdiagnosed as HCC, surgery is the choice of treatment for these patients. Although their pathology resembles malignant lymphoma, the clinical course is completely benign. The authors propose that RLH of the liver can be discriminated from HCC by its clinical features.

Cytomegalovirus frequency in neonatal intrahepatic cholestasis determined by serology, histology, immunohistochemistry and PCR

AIM: To determine cytomegalovirus (CMV) frequency in neonatal intrahepatic cholestasis by serology, histological revision (searching for cytomegalic cells), immunohistochemistry, and polymerase chain reaction (PCR), and to verify the relationships among these methods.METHODS: The study comprised 101 non-consecutive infants submitted for hepatic biopsy between March 1982 and December 2005. Serological results were obtained from the patient's f iles and the other methods were performed on paraffin-embedded liver samples from hepatic biopsies. The following statistical measures were calculated: frequency, sensibility, specif ic positive predictive value, negative predictive value, and accuracy.RESULTS: The frequencies of positive results were as follows: serology, 7/64 (11%); histological revision, 0/84; immunohistochemistry, 1/44 (2%), and PCR, 6/77 (8%). Only one patient had positive immunohistochemical fi ndings and a positive PCR. The following statistical measures were calculated between PCR and serology: sensitivity, 33.3%; specificity, 88.89%; positive predictive value, 28.57%; negative predictive value, 90.91%; and accuracy, 82.35%. CONCLUSION: The frequency of positive CMV varied among the tests. Serology presented the highest positive frequency. When compared to PCR, the sensitivity and positive predictive value of serology were low.

Isolated pancreatic granulocytic sarcoma: A case report and review of the literature

Granulocytic sarcoma (GS) is an extramedullary tumor mass consisting of immature myeloid cells. Isolated pancreatic granulocyte sarcoma is extremely rare. We report a very unusual pancreatic granulocytic sarcoma in a patient without acute myeloid leukemia. The patient presented with acute epigastric pain because of splenic infarction due to a mass consisting of myeloblasts in the pancreatic tail. The patients underwent splenectomy and distal pancreatectomy. Pathology and immunohistochemistry suggested a GS. Despite local surgery, an isolated tumor recurred 2 mo after operation and the patient died 3 mo after removal of the tumor. Only 7 reported cases of pancreatic GS were identified in the literature and the mass was located in the pancreatic head. This is the first report of GS in the pancreatic tail with splenic infarction.