鼻咽癌转移相关的分泌蛋白质的筛选

目的：筛选鼻咽癌（nasopharyngeal carcinoma, NPC）转移相关的分泌蛋白质,为阐明NPC转移机制以及筛选NPC转移分子标志物提供实验依据。方法：应用二维凝胶电泳（two-dimensional electrophoresis,2-DE）技术分离一对来自同一亲本,具有不同转移潜能的NPC细胞系5-8F和6-10B细胞的分泌蛋白质,图像分析识别差异表达的蛋白质点,基质辅助激光解吸电离飞行时间质谱(MALDI-TOF-MS)和电喷雾-四极杆-串联质谱(ESI-Q-TOF MS/MS)对差异表达的蛋白质点进行鉴定。Western印迹法检测差异分泌蛋白质nm23-H1在两株细胞中的表达水平。结果：建立了5-8F和6-10B分泌蛋白质的2-DE图谱,质谱分析鉴定出14个非冗余的分泌蛋白质,其中Oncoprotein18等蛋白质在高转移潜能NPC细胞系5-8F中的表达水平高于无转移潜能的NPC细胞系,而nm23-H1等蛋白质在5-8F细胞系中的表达水平低于6-10B细胞系。Western印迹分析证实了nm23-H1在5-8F和6-10B细胞分泌蛋白质中的差异表达水平。结论：所鉴定的14个非冗余的差异分泌蛋白质为研究NPC转移机制以及筛选NPC转移分子标志物提供了实验依据。     

鼻咽癌组织与正常鼻咽上皮组织的差异磷酸化蛋白质组分析

目的：比较鼻咽癌(nasopharyngeal carcinoma, NPC)组织与正常鼻咽上皮组织磷酸化蛋白质组差异,筛选差异磷酸化蛋白质,为揭示NPC的发病机制提供依据。方法：采用双向凝胶电泳技术(2-DE)分离NPC组织与正常鼻咽上皮组织的总蛋白质,蛋白质转膜后与抗酪氨酸磷酸化抗体进行Western印迹分析,图像分析识别差异磷酸化蛋白质点,电喷雾-四极杆-串联质谱（ESI-Q-TOF MS/MS）鉴定差异的酪氨酸磷酸化蛋白质,采用NetPhos软件预测蛋白质的酪氨酸磷酸化位点,并采用生物信息学方法对差异磷酸化蛋白质的功能和亚细胞定位进行分析。采用Western印迹检测差异蛋白质（phosphatidylethanolamine- binding protein 1）在正常鼻咽黏膜组织和鼻咽癌组织的总蛋白质中的磷酸化水平。结果：建立了NPC组织与正常鼻咽上皮组织的酪氨酸磷酸化蛋白质表达谱,识别了25个差异酪氨酸磷酸化蛋白质,共鉴定了13个差异酪氨酸磷酸化蛋白质,其中7个蛋白质的酪氨酸磷酸化水平在NPC组织中增强,而6个蛋白质的酪氨酸磷酸化水平降低。NetPhos软件预测和生物信息学分析结果显示,13个差异蛋白质均存在酪氨酸磷酸化位点,其功能涉及物质代谢、细胞结构、细胞防御以及信号转导。与正常鼻咽黏膜组织相比较,差异蛋白phosphatidylethanolamine-binding protein 1在鼻咽癌组织中磷酸化表达水平下调。结论： 鉴定了13个可能与NPC发病相关的酪氨酸磷酸化蛋白质,为揭示NPC发病机制提供了科学论依据。     

5-aza-2-dC处理鼻咽癌细胞前后的差异蛋白质组学研究

目的：比较5-杂氮-2′-脱氧胞苷（5-aza-2-dC）处理鼻咽癌细胞前后蛋白质组的差异,为筛选鼻咽癌的甲基化失活基因提供依据。方法：用去甲基化药物5-aza-2-dC处理鼻咽癌细胞系5-8F细胞,应用双向凝胶电泳（2-DE）技术分离5-aza-2-dC处理与未处理5-8F细胞的蛋白质,PDQuest图像分析软件识别药物处理与未处理5-8F细胞的差异表达蛋白质点,基质辅助激光解吸电离飞行时间质谱（MALDI-TOF-MS）鉴定差异表达的蛋白质。Western 印迹法检测差异蛋白质nm23-H1在药物处理与未处理5-8F细胞中的表达水平。结果：建立了5-aza-2-dC处理与未处理5-8F细胞蛋白质的2-DE图谱,识别了49个差异表达的蛋白质点,鉴定了33个差异表达的蛋白质,其中15个蛋白质在5-aza-2-dC处理5-8F后表达上调。Western 印迹分析证实了nm23-H1在药物处理与未处理5-8F细胞中的差异表达水平。结论：15个5-aza-2-dC处理后表达上调蛋白质的编码基因可能是5-8F细胞的甲基化沉默基因。     

克罗恩病患者与健康人血清蛋白质组学比较初步研究

目的：应用蛋白质组学方法在患者血清中寻找与克罗恩病相关的蛋白质。方法: 采取克罗恩病患者以及正常成人血清蛋白样本各4例，用不同的CyDye荧光染料标记后进行胶内差异双向凝胶电泳(2-D DIGE)，并对获得的图谱进行分析及对差异蛋白质进行基质辅助激光解吸离子化飞行时间质谱(MALDI-TOF-MS) 鉴定和生物学信息分析。结果: 通过2-D DIGE分析，发现了克罗恩病患者中存在29个表达异常蛋白质点，质谱分析和数据库检索共鉴定出22种蛋白质，包括SER/THR，CD45，APC等。结论: 蛋白质组学能很好显示克罗恩病患者与正常人血清蛋白质表达差异，本研究鉴定的蛋白质可能为研究克罗恩病的生物学行为提供新的分子标记物。     

人膝骨关节炎血清生物学标志物的筛选

目的: 比较膝骨关节炎患者与正常人血清蛋白质组的差异,筛选其能用于骨关节炎诊断、治疗或发病机制研究的血清生物学标志物。方法: 利用双向荧光差异凝胶电泳技术比较膝骨关节炎患者血清(4例)与健康志愿者血清(4例)蛋白图谱的差异,使用基质辅助激光解吸电离-飞行时间质谱及生物信息学相关技术对获得的差异蛋白进行鉴定和初步分析,并用Western blotting进一步验证所鉴定的蛋白。结果: 成功建立了骨关节炎血清差异蛋白质组学的研究方法;通过质谱分析初步鉴定出8个差异蛋白,其中5个蛋白在膝骨关节炎组表达上调,3个蛋白在膝骨关节炎组表达下调。Western blotting进一步验证得到α₂-巨球蛋白在膝骨关节炎组表达上调,与双向荧光胶内差异凝胶电泳技术联合质谱分析的结果一致。结论: 膝骨关节炎患者血清与正常人血清蛋白表达存在明显差异,其中α₂-巨球蛋白可作为骨关节炎潜在的疾病相关生物学标志物进一步研究,为骨关节炎的诊断、治疗和发病机制的研究提供新的线索和实验基础。     

蛋白质组学技术分析5-aza-2dC处理白?∠赴鸋L-60前后的差异表达蛋白质

目的：研究甲基化转移酶抑制剂5-杂氮-2’-脱氧胞苷(5-aza-2’-deoxycitydine, 5-aza-2dC)对人急性髓系白血病细胞HL-60细胞蛋白质表达谱的影响，探讨5-aza-2dC抗急性髓系白血病作用机制。方法：采用二维凝胶电泳（2-DE)技术分离5-aza-2-dC处理与未处理HL-60细胞的总蛋白质，PDquest 图像分析软件识别差异蛋白质点，基质辅助激光解吸电离飞行时间质谱(MALDI-TOF-MS)鉴定差异蛋白质点。 然后采用Western 印迹检测差异蛋白质半乳糖凝集素1（Galectin-1） 在药物处理与未处理HL-60细胞中的表达水平。结果： 建立了5-aza-2dC 处理与未处理HL-60 细胞蛋白质的2-DE 图谱，识别了35个差异表达的蛋白质点，鉴定了27个差异表达的蛋白质，其中包括Galectin-1在内的21个蛋白质在5-aza-2dC 处理后的HL-60 细胞中表达上调，6个蛋白质表达下调或缺失。Western 印迹结果证实Galectin-1在5-aza-2dC 处理HL-60细胞中表达上调。结论：21个表达上调蛋白质的编码基因可能是HL-60细胞的甲基化失活基因，它们可能与急性髓系白血病发病以及5-aza-2-dC抗人急性髓系白血病细胞作用有关     

系统性红斑狼疮肾炎血清差异蛋白质的筛选

目的:对比分析系统性红斑狼疮肾炎(LN)患者和健康人双重滤过血浆置换(DFPP)产物中的差异蛋白质,筛选与LN发生相关的血清蛋白质。方法:利用DFPP的方法分别对4例LN患者和4例健康人血浆进行过滤,将滤下的大分子蛋白混合物行双向电泳分离并银染显影,经图像分析筛选差异表达的蛋白点。应用QSTAR Pulsar I型串连飞行质谱鉴定差异表达的蛋白点,通过生物信息学比对,对其中有意义分子行血清浓度检测验证。结果:2组间筛选出差异蛋白点共317个,其中表达量差异达3倍以上的蛋白点69个,经串连质谱分析鉴定出包括人冷凝集素IgM抗体、免疫球蛋白λ轻链、谷胱甘肽-S-转移酶、结合珠蛋白前原蛋白、甲状腺素转运蛋白、载脂蛋白E、皮质类固醇结合球蛋白、玻连蛋白、角蛋白和转铁蛋白等十多个在SLE活动性肾炎血清中表达改变的蛋白分子。ELISA及RIA检测结果与双向电泳相符。结论:LN患者血清与健康人血清的蛋白表达谱有一定差异,这些差异分子可能成为LN诊断及预后判断的重要标志物。     

蛋白质组学检测β-原肌球蛋白在结肠癌组织中的表达

目的：通过比较结肠腺癌与正常结肠黏膜的蛋白质组表达差异,寻找与结肠腺癌发生相关的蛋白质,筛选结肠癌诊断的分子标志物。方法: 运用蛋白质组学技术,对8例结肠腺癌组织和8例正常结肠黏膜组织进行胶内差异双向电泳（2-D）,选择差异表达超过2倍的蛋白质进行MALDI-TOF/TOF质谱分析和生物信息学分析,并对结肠癌组织中表达下调的蛋白β-原肌球蛋白(TM β)进行Western blotting验证。结果: 成功建立结肠腺癌和正常结肠黏膜的双向凝胶电泳图谱,结肠腺癌和正常黏膜组织凝胶电泳图谱中平均蛋白质斑点数分别为3 289和3 066,其中表达差异超过2倍的斑点共有31个,质谱分析和数据库检索共鉴定出18种蛋白质,包括cytokeratin 8、cytokeratin 10、S100A6、TM β、protein disulfide isomerase 等。从功能分析,这些差异蛋白与癌细胞的发生、增殖、分化、转移等相关;TM β在结肠癌组织中的表达水平Western blotting结果与电泳结果一致。结论: 蛋白质组学能有效地分离和鉴定结肠腺癌组织与正常结肠组织间的差异表达蛋白质,结肠癌组织中表达下降的TMβ,可能成为结肠癌诊断的分子标记物。     

急性期神经根型颈椎病患者的血清差异蛋白组学分析

背景：正常健康人向急性期神经根型颈椎病转化的具体分子机制尚未完全明确，需进一步深入研究。目的：探讨正常健康人群与急性期神经根型颈椎病患者的血清蛋白组学差异表达，并寻找及鉴定两者之间的潜在特异性血清标志物。方法：收集急性期神经根型颈椎病患者、正常健康人群血清各8例，应用绝对定量技术联合液相色谱-串联质谱技术进行蛋白组学筛选与分析，以期探寻与鉴定急性期神经根型颈椎病患者差异表达的血清蛋白质。结果与结论：(1)绝对定量技术筛查出有意义差异蛋白183种，鉴定出11种显著性差异蛋白质(P <0.05)，其中与正常健康人组相比，急性期神经根型颈椎病组中人类白细胞抗原A、分泌珠蛋白家族1a成员1、蛋白4-羟基苯丙酮酸双加氧酶等3种差异蛋白显著上调，免疫球蛋白IgG3恒定区、皮肤因子、肌球蛋白轻链3等7种差异蛋白显著下调；(2)基因本体富集分析结果表明，这些差异蛋白参与了抗原结合、免疫球蛋白受体结合等分子功能；(3)蛋白相互作用网络分析表明，正常健康人群与急性期神经根型颈椎病之间的共同差异蛋白中HLA-A、HPD、PSMA3、DMKN、SCGB1A1、MYL3等6种位于功能网络节点，且与机体免疫...     

鲍曼不动杆菌脓毒症大鼠中性粒细胞的蛋白质组学研究

目的 通过分离并鉴定鲍曼不动杆菌脓毒症大鼠中性粒细胞的差异表达蛋白质,以发现可能的在鲍曼不动杆菌脓毒症中起关键作用的蛋白和可用于早期诊断的鲍曼不动杆菌脓毒症标志物.方法 提取正常大鼠中性粒细胞和鲍曼不动杆菌脓毒症大鼠中性粒细胞的总蛋白质,用双向凝胶电泳分离蛋白并进行比较.选择在鲍曼不动杆菌脓毒症中明显差异表达的蛋白点,行质谱分析,并选取蛋白加以免疫印迹验证.结果 获得了分辨率和重复性均很好的凝胶蛋白图谱.筛选出的在鲍曼不动杆菌脓毒症中明显差异表达的50个蛋白点,共有41个蛋白点被成功鉴定,其中在鲍曼不动杆菌脓毒症中高表达的为24个,低表达的为17个.Western blot蛋白验证基本与质谱鉴定一致.结论 鲍曼不动杆菌脓毒症大鼠中性粒细胞相对于正常大鼠中性粒细胞蛋白存在明显的差异,通过蛋白质组学方法筛选并鉴定出的这些蛋白质可能会成为用于鲍曼不动杆菌脓毒症早期诊断和治疗的分子靶点.     

Proteomic identification of alpha-2-HS-glycoprotein as a plasma biomarker of hypopharyngeal squamous cell carcinoma

Hypopharyngeal squamous cell carcinoma (HSCC) has very poor prognosis compared with other head and neck squamous cell carcinomas. Late-stage diagnosis of HSCC increases mortality. Therefore, more effective biomarkers for early diagnosis of HSCC are necessary. Unfortunately, appropriate biomarkers for clinical diagnosis and prognosis have not been identified yet. However, recent progresses in quantitative proteomics have offered opportunities to identify plasma proteins as biomarkers for HSCC. In the present study, plasma samples were analyzed by two-dimensional differential gel electrophoresis (2D-DIGE), and differentially expressed proteins were identified by matrix assisted laser desorption ionization-time of flight/time of flight mass spectrometry (MALDI-TOF/TOF MS). A total of 26 proteins representing 12 unique gene products were identified. The up-regulation proteins were alpha-2-HS-glycoprotein (AHSG), complement C4-B, haptoglobin, C-reactive protein, and ceruloplasmin, whereas the down-regulation proteins were serum albumin, angiotensinogen, alpha-1-antichymotrypsin, Ig gamma-3 chain C region, fibrinogen gamma chain, apolipoprotein A-I, and Ig kappa chain C region. Among all the differentially expressed proteins, AHSG was validated by western blot and ELISA. The results were consistent with the data from 2D-DIGE, further suggesting that AHSG may be employed as a potential biomarker for the early diagnosis of HSCC. In summary, this study was the first to use 2D-DIGE and MALDI-TOF/TOF platform to identify the potential plasma biomarkers for HSCC. The plasma AHSG showed great potential for HSCC screening.     

Proteome analysis of autoantibodies in sera of patients with cancer

Circulating autoantibodies are useful diagnostic markers of cancers and autoimmune diseases. Research over the past decade has resulted in some reports on the presence of autoantibodies against disease-related proteins such as annexin-I & II, recoverin and protein gene product 9.5 in the sera of patients with lung cancer, and also against calreticulin and alpha-enolase in autoimmune diseases. In this study, we first identified the a-enolase autoantibody in the sera of patients with lung adenocarcinoma by proteomics-based analysis. The comparison of two-dimensional polyacrylamide gel electrophoresis (2D-PAGE)/western blot (WB)/ECL detection revealed distinct distributions of antibodies in the sera of lung adenocarcinoma, tuberculosis and healthy subjects which reacted with soluble proteins derived from the adenocarcinoma A549 cell line. We found 16 spots in patients with adenocarcinoma by 2D-PAGE/WB/ECL detection and identified alpha-enolase, chaperonin, and other autoantibodies in the adenocarcinoma patients' sera. The specificities of an antibody against alpha-enolase was preliminarily observed in sera from 3 of 5 patients with adenocarcinoma, 0 of 10 patients with tuberculosis and 0 of 10 healthy subjects. In conclusion, we first identified alpha-enolase autoantibody in sera of lung adenocarcinoma and the autoantibody was seemed to be a specific marker of the lung adenocarcinoma. In addition, we also identified various autoantibodies in esophageal cancer, hepatocellular carcinoma, and non-Hodgkin's lymphoma. Moreover, we tried to identify the corresponding antigen of an unknown anti-cytoplasmic autoantibody, and an anti-red blood cell antibody by proteomics-based analysis. These antibodies might become new diagnosis markers.     

Characterization of immunoglobulin G and its subclass response to Indian kala-azar infection before and after chemotherapy

Serologic parameters of kala-azar were evaluated by Western blot analysis. Sera from kala-azar patients with confirmed diagnoses were screened for immunoglobulin G (IgG) and IgG subclass-specific reactivity against Leishmania donovani membrane antigen (LAg). Heterogeneous LAg-specific IgG reactivity with numerous proteins with molecular masses ranging from 18 to 190 kDa was observed. Though the individual band patterns were varied, seven polypeptides of approximately 31, 34, 51, 63, 72, 91, and 120 kDa were immunoreactive with all the sera tested from kala-azar patients. The band patterns of the immunoblots of sera from patients after treatment and clinical cure with sodium antimony gluconate revealed a decrease in the frequency of the bands. Still, recognition of the 63- and 120-kDa bands was 100%, and the 55- and 91-kDa fractions were recognized in 93% of the sera from cured individuals. Among the IgG subclasses, IgG1 reacted with the greatest number of polypeptides. The 63-kDa protein was again detected by all of the IgG subclasses of all the sera tested. Other fractions recognized by the subclasses of more than 70% of the serum samples included those of 47, 51, 55, and 78 kDa. Following treatment, 63- and 51-kDa bands were the most reactive with the IgG subclasses. LAg-associated cross-reaction with other reference human antisera revealed a mild reactivity of the 63-kDa polypeptide with some of the serum samples from leprosy, malaria, typhoid, tuberculosis, and healthy controls. Western blot analysis of LAg entrapped in liposomes, strong vaccine candidates against experimental visceral leishmaniasis, revealed a more restricted band pattern. The 63-kDa fraction revealed by all pre- and posttreatment sera showed almost negligible levels of cross-reaction with sera from patients with other diseases or from healthy controls. These observations provide insight into induced immunity during kala-azar infection for future application.     

Elevated expression of complement C3 protein in chemically induced hepatotumorogenesis in Wistar rats: A correlative proteomics and histopathological study

Liver cancer remains the leading cause of cancer-related mortality worldwide. Early detection of liver cancer is problematic due to the lack of a marker with high diagnosis sensitivity and specificity. The present study was designed to determine the differently expressed proteins at early stage in the serum of animals with liver cancer vis-à-vis controls and figure out the function of the proteins. One-dimensional electrophoresis (1D), two-dimensional electrophoresis (2DE) and liquid chromatography mass spectrometry (LC–MS/MS) were used to screen the serum proteins of liver cancer induced in animals by diethyl nitrosamine (DEN) + 2-acetyl amino fluorine (2-AAF). From optimized 2DE image and computer assisted PD Quest analysis were found to be differentially expressed spots when the serum from normal and treated animals were compared. Among these, one spot was selected whose expression level was higher in DEN + 2-AAF treated animal sera than in adjacent normal animal sera. The target spot was excised from the 2D gel of liver cancer sera and the peptide mass fingerprinting as obtained LC–MS/MS analysis after digesting the chosen protein spot. This was identified to be complement C3 protein. The changes in complement C3 expression level were validated by Western blot analysis. We reported that the changes in complement C3 concentration start at very early stage of tumorogenesis. The fully grown tumors were developed at 120 days and hepatotumorogenesis was confirmed by histopathological examination. This protein may therefore represent a powerful tool in search for candidate biomarkers for HCC.     

Cockroach-allergen study: allergen patterns of three common cockroach species probed by allergic sera collected in two cities

Antigens/allergens of three common cockroach extracts, crude whole body extract of the American cockroach (CRa-A), crude whole body extract of the German cockroach (CRa-G), and crude whole body extract of the Oriental cockroach (CRa-O), were studied with crossed immunoelectrophoresis, crossed radioimmunoelectrophoresis, and Western blot analysis. Sera of cockroach-allergic patients with asthma, 10 from Chicago, Ill. (C group) and six patients from Lexington, Ky. (L group), were used; results were then compared with sera of control subjects with asthma. Qualitative differences in protein bands were noted among CRa-A, CRa-G, and CRa-O by crossed immunoelectrophoresis and sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Allergen bands on Western blot were analyzed for distribution by molecular weight (MW) with relative intensity scores. Results were compared by species and by geography. Two to 12 allergenic bands of variable MW (14 kd to greater than 116 kd) were identified by 13 of 16 individual sera from cockroach-allergic patients from all three extracts. CRa-A demonstrated 55 bands with an intensity score of 125; CRa-G, 58 bands with an intensity score of 100; and CRa-O, 51 bands with an intensity score of 108. Allergenic bands of CRa-A were identified by six sera of the C group and one sera of the L group, whereas bands of both CRa-G and CRa-O were noted by nine sera of the C group and four sera of the L group. All three species had an allergen band in MW range of 40 to 45 kd that reacted to most sera from cockroach-allergic patients with asthma.     

Human antibody response to human cytomegalovirus-specific DNA-binding proteins

Human antibody responses to human cytomegalovirus (HCMV) specific DNA-binding proteins were studied in serum samples by the Western blot technique. The molecular weights of six DNA-binding proteins found in HCMV-infected cells, ranged from 52kD to 18kD. The sera obtained from patients with acute HCMV infections reacted well with the six HCMV specific DNA-binding proteins. The strongest reactivity was observed with the 52kD and 35kD proteins. The sera from healthy HCMV seropositive donors reacted only with the 52kD DNA-binding protein as visualized in Western blots, but 2 out of 8 sera failed to react with any HCMV specific DNA-binding proteins.     

The assessment of anti-endothelial cell antibodies in scleroderma-associated pulmonary fibrosis. A study of indirect immunofluorescent and western blot analysis in 49 patients with scleroderma

We recently reported on the use of an indirect immunofluorescent method designated the rodent lung assay; this test assesses for the presence of circulating antibodies directed at components of the microvasculature. Serum samples from 49 patients with scleroderma were incubated with rodent lung tissue sections and visualized with fluoresceinated human anti-IgG. The assay also was performed on samples from a control group. Western blot analysis was performed with endothelial cell protein extracts using serum samples from patients with scleroderma and from healthy control subjects. The control subjects had a negative indirect immunofluorescent assay result. In the patients with scleroderma, there was a significant positive correlation between intensity of indirect immunofluorescent staining and pulmonary fibrosis (r = 0.316; P = .0347) and hypertension (r = 0.310; P = .0408). Western blot analysis revealed antibody binding to proteins in extracts of human endothelial cells in all patients in whom there was evidence of pulmonary disease. The indirect immunofluorescent rodent lung assay and Western blot data support a potential role of anti-endothelial cell antibodies in the propagation of scleroderma-associated pulmonary disease.     

G-protein coupled receptor-associated sorting protein 1 (GASP-1), a potential biomarker in breast cancer

An innovative “2-D high performance liquid electrophoresis” (2-D HPLE) technology was used to identify serum biomarkers associated with the early stage of breast cancer in addition to other more advanced stages. 2-D HPLE is a newly developed electrophoretic technology that separates 100s of serum albumin complexes on a polyvinyl membrane based on their surface charges. Association of cancer proteins or their fragments (biomarkers) with pre-existing albumin complexes in the blood of cancer patients results in altered mobility on the membrane. Using 2-D HPLE we identified that a specific fragment of G-protein coupled receptor-associated sorting protein 1 (GASP-1) was present in the sera of patients with early stage disease but absent in sera of normal patients. GASP-1 has been shown to modulate lysosomal sorting and functional down-regulation of a variety of G-protein coupled receptors in neuronal cells. However, no reports have linked GASP-1 to breast cancer pathogenesis. GASP-1 was detected in tumor extracts of 7 cases of Stage 2 and Stage 3 breast cancers, but not in adjacent normal tissue as revealed by western blot analysis using an antibody developed against a GASP-1 peptide identified by our 2-D HPLE technology. Using this antibody, we immunohistochemically detected over-expression of GASP-1 in all of 107 cases of archived ductal breast carcinoma tumor samples, while normal adjacent breast tissue from 12 cases of ductal carcinoma showed little or no staining. Additionally, all 10 cases of metastatic breast carcinoma present in lymph nodes were positive. Strong positive GASP-1 staining was observed in all tumor tissue including ductal carcinoma in situ (DCIS) and invasive ductal carcinoma. Additionally, we observed a wide spectrum of enhanced staining of premalignant ductal epithelial cells present in benign ducts and those found in atypical ductal hyperplasia (ADH). These studies identify GASP-1 as a potential new serum and tumor biomarker for breast cancer and suggest that GASP-1 may be a novel target for the development of breast cancer therapeutics.