抗人铁蛋白单克隆抗体的制备及其在肝癌病人血清铁蛋…

铁蛋白是一种肿瘤相关蛋白质,我们从人肝癌组织中分离纯化了铁蛋白,并筛选到一株抗该铁蛋白的单克隆抗体杂交瘤细胞株（６Ｄ６）,它针对铁蛋白上的构象决定簇,并对人源铁蛋白高度专一。然后我们用此单抗建立了测定铁蛋白浓度的夹心ＥＬＩＳＡ法,并对方法的灵敏度,精确度及特异性作了研究。本法用于测定不同血清标本的结果表明：肝癌病人血清中的铁蛋白浓度明显高于正常人,这可能对临床诊断会有使用价值。     

单抗在人铁蛋白免疫亲和纯化中的应用

抗人铁蛋白单抗６Ｄ６和Ａ－ｈＦ－Ｃ与肝型铁蛋白和心型铁蛋白的反应性有所不同。６Ｄ６对两种铁蛋白的反应性相似;而Ａ－ｈＦ－Ｃ单抗与肝型铁蛋白的反应性较强。我们将６Ｄ６和Ａ－ｈＦ－Ｃ分别制成了亲和凝胶,用来纯化人肝脏和心脏粗抽提物中的铁蛋白。此法具有操作简便,产率和产品纯度高等优点。     

人心肌酸性铁蛋白的分离纯化及临床应用

人心肌匀浆经热变性,酸化,硫酸铵盐析,超离心、Ｓｅｐｈａｒｏｓｅ ＣＬ－４Ｂ柱层析和制备等电聚焦分离,得到酸性铁蛋白,经鉴定,所得酸性铁蛋白ｐＩ为５．０,Ｈ亚基分子量为２１ｋＤ,Ｌ亚基为１９ＫＤ,ＰＡＧＥ分析呈单一区带,制备了兔抗人酸性铁蛋白抗血清,用该抗血清建立的人酸性铁蛋白放射免疫分析可检测出８０％甲胎蛋白阴性肝癌病人。     

人心肌酸性铁蛋白的分离纯化及临床应用

人心肌匀浆经热变性、酸化、硫酸铵盐析、超离心、ＳｅｐｈａｒｏｓｅＣＬ－４Ｂ柱层析和制备等电聚焦分离,得到酸性铁蛋白。经鉴定,所得酸性铁蛋白ｐＩ为５．０,Ｈ亚基分子量为２１ｋＤ,Ｌ亚基为１９ｋＤ,ＰＡＧＥ分析呈单一区带。制备了兔抗人酸性铁蛋白抗血清,用该抗血清建立的人酸性铁蛋白放射免疫分析可检测出８０％甲胎蛋白阴性肝癌病人。     

铁蛋白分析技术的研究进展

铁蛋白是含铁蛋白质,1943年Cranick用硫酸铵沉淀、超滤、硫酸镉重结晶、凝胶层析等技术,获得铁蛋白,经免疫电泳、等电聚焦电泳和离子交换层析鉴定,血清铁蛋白由含铁单体,     

铁蛋白结构和功能的生物信息学分析

[目的]通过生物信息学方法探究铁蛋白的结构和功能的相关信息,研究铁蛋白与肝癌的潜在关系。[方法]从NCBI获取铁蛋白序列,分别使用ProScale、TMHMM、NCBI-CDD和Prosite工具分析铁蛋白的亲疏水性、跨膜区、功能结构域和生物活性位点。使用SOPMA、PSIPRED和SWISS-MODEL平台预测了铁蛋白的二级结构和三级结构。进一步,基于STRING数据库研究了铁蛋白的互作蛋白和富集通路。[结果]铁蛋白是一种亲水性蛋白,无跨膜区,存在1个铁蛋白样二铁氧化的功能结构域和2个生物活性位点,具有较高的螺旋程度。铁蛋白与RPS17、TFRC、SCARA5、NCOA4和COX8A五种蛋白相互作用,这些蛋白显著富集在铁死亡与矿物吸收富集途径,与癌症的发生有关。[结论]对铁蛋白进行生物信息学分析,预测了铁蛋白重链和轻链的理化性质和空间结构,也从其功能上揭示了与肝癌的潜在关系,为深入研究提供了理论基础。     

铁蛋白的生物工程应用

铁蛋白是一种普遍存在于生物体内的储铁蛋白,具有铁离子代谢、抗氧化胁迫及消除其他过量金属离子毒害作用的功能。随着对铁蛋白结构和生化功能认识的深入,铁蛋白作为一个含有四氧化三铁核心的特殊蛋白复合体,被广泛应用于生物医学、纳米材料、生物分子成像等各种生物工程领域。该综述针对已知的主要铁蛋白分子,论述了铁蛋白的结构及酶活性机理,基于铁蛋白的多功能分子骨架应用,以及基于铁蛋白磁性的生物分子开关等热点研究,最后对铁蛋白生物工程、生物医学领域的应用和发展进行了展望。     

西安地区妊娠妇女不同孕期血清铁蛋白、叶酸和维生素B_(12)的水平及临床意义

目的:探讨西安地区妊娠妇女不同孕期血清铁蛋白、叶酸和维生素B12的水平及临床意义。方法:收集2012年8月至2013年1月在本院进行产前检查的750例妊娠期妇女及86例健康非妊娠妇女的血清标本。采用全自动化学发光免疫分析法测定其血清铁蛋白、叶酸和维生素B12的含量。结果:孕妇血清铁蛋白、叶酸和维生素B12的含量随孕期的增长而逐渐降低。早、中、晚孕期妇女叶酸的含量较对照组相比,并无显著差异(P0.05)。中、晚孕期妇女血清铁蛋白和维生素B12含量均显著低于对照组,差异有统计学意义(P0.05)。结论:西安地区中、晚孕期的孕妇体内存在铁蛋白和维生素B12缺乏,但并不缺乏叶酸。在重视补充叶酸的同时,也应该重视对铁和维生素B12的补充。     

乳铁蛋白与骨质疏松症

本文主要阐述了乳铁蛋白对造骨细胞、破骨细胞及半胱氨酸蛋白酶的作用,由于其对造骨细胞有促进作用及对破骨细胞和半胱氨酸蛋白酶（如K、L）的抑制作用,因此乳铁蛋白是治疗骨质疏松症的一种潜在的治疗手段。     

动物储铁蛋白研究进展

铁元素是生物体中必不可少的微量元素,在生物的生长发育中发挥着重要作用。铁蛋白是一种分布广泛的球形蛋白,能够以稳定的形式储存大量铁。铁蛋白通过储存和释放铁来维持机体内铁平衡。铁蛋白不仅是机体中重要的铁储存蛋白,同时也能有效保护生物体免受来自氧自由基的损伤。与此同时,铁蛋白含量可以作为一些疾病预防检测的明确指标。对铁的代谢吸收及铁对基因调控的研究,进一步说明了维持铁平衡对生物体有重要意义。     

Properties of human tissue isoferritins.

1. Human liver ferritin was separated by preparative isoelectric focusing into six fractions. 2. Except for the least acidic fraction the reactivity with antibody against spleen ferritin increased with rising pI, but with antibody against heart ferritin the reactivity decreased. 3. The highest iron content was found in the most acidic isoferritins and progressively decreased with rising pI. 4. Iron uptake was studied in apoferritin prepared from heart and liver ferritin fractions separated by ion-exchange chromatography. There was good correlation between the rate of iron uptake and pI. The most acidic fractions took up iron more rapidly than did the more basic ones. 5. Ferritin was prepared from heart, liver, spleen and kidney. There was little difference on isoelectric focusing between ferritin obtained from normal tissues and the corresponding iron-loaded tissues from patients who had received multiple blood transfusions. The iron-loaked heart ferritin invariably contained relatively more of the basic isoferritins. Normal and iron-overloaded heart ferritins were separated into isoferritin fractions by ion-exchange chromatography, and in each case there was a fall in iron content as the pI increased. The iron content of ferritin from the iron-overloaded heart was higher throughout than that from normal heart. 6. There is a relationship between the rate of iron uptake by apoferritin and pI, and this probably accounts for the variation in iron content of the isoferritins found in human liver and heart.     

Isolation of ferritin from human mammary and pancreatic carcinomas by means of antibody immunoadsorbents

In the course of a study of tumor antigens we prepared an absorbed antiserum to a breast tumor that reacted strongly with breast tumor but not with normal breast. The antigen was purified by adsorption to an antibody immunoadsorbent prepared from this antiserum, elution with 2 m potassium thiocyanate at neutral pH, and passage through an immunoadsorbent containing antibodies to human serum. The purified antigen was identified as ferritin by electrophoretic, chemical, and immunological criteria. Isoelectric focusing in acrylamide gels revealed that tumor ferritin contained six bands seen in normal liver ferritin plus a variable number of acidic components not detected in normal liver ferritin. The acidic components were concentrated by chromatography on a DEAE-cellulose column. Similar acidic components described previously in ferritins isolated from cultured human tumor cells, hepatomas, and fetal liver have been designated as “carcino-fetal” ferritins.     

Applications of a monoclonal antibody to human ferritin in various immunoassays

Monoclonal antibodies to human liver ferritin were generated by an improved hybridoma technique using a semisolid medium containing methylcellulose for initial cloning after the cell fusion. Out of more than 1000 hybrid clones, only 1 was shown to secrete high-affinity monoclonal antibodies to human liver ferritin. The immunoglobulin subclass of this antibody was determined to be IgG2. The association constant between liver ferritin and this antibody was determined to be greater than 1 X 10(10) M-1. Due to the oligomeric nature of ferritin, this antibody can be simultaneously utilized as the first and second antibody in solid-phase sandwich immunoradiometric and enzyme immunoassays. This immunoassay procedure can be performed within 30-45 min and has a sensitivity of about 1 ng/ml. Under identical assay conditions, ferritin isolated from human spleen and human heart gave 50 and 30% cross-reactivity, respectively.     

Mössbauer spectroscopy of the iron cores in human liver ferritin, ferritin in normal human spleen and ferritin in spleen from patient with primary myelofibrosis: preliminary results of comparative analysis

Comparative study of human liver ferritin and spleen tissues from healthy human and patient with primary myelofibrosis was carried out using Mössbauer spectroscopy with a high velocity resolution at 295 and 90 K and with a low velocity resolution at 20 K. The results obtained demonstrated that the iron content in patient’s spleen in the form of iron storage proteins was about ten times larger than that in normal tissue. However, in the case of patient with primary myelofibrosis the magnetic anisotropy energy barrier differed from that in normal case and, probably, the iron core size was supposed to be slightly larger than that in both normal spleen tissue and normal human liver ferritin in contrast to well-known data for iron overload in patients with thalassemia accompanied by the iron-core size increase. Therefore, the iron overload in the case of patient with primary myelofibrosis may be related to increase in the ferritin content mainly. It was also found that Mössbauer hyperfine parameters for normal and patient’s spleen and normal human liver ferritin demonstrated some small differences related, probably, to some small structural variations in the ferritin iron cores of patient’s spleen.     

Alteration in tryptic peptide patterns of ferritins purified from human colon carcinoma

Ferritin from malignant tissue differs electrophoretically from normal ferritin. The molecular basis of this difference has not yet been defined. Malignant tissue contains a mixture of ferritins from normal cells, inflammatory cells as well as cancer cells. GW-39 is a pure colon carcinoma cell system that synthesizes human carcinoembryonic antigen. Therefore, ferritin was isolated from normal colon mucosa and colon cancer tissues, as well as from the colon carcinoma cell line, to clarify the molecular relationship between normal and malignant ferritins. Colon carcinoma ferritin differs in primary structure from normal colon mucosal ferritin and contains at least six additional different tryptic peptides. These six peptides were also found in the ferritin from the colon carcinoma cell line. These data suggest that the alteration in ferritin structure occurs at the cellular level and is associated with the malignant state.     

内蒙地区发热患者血清中冠状病毒抗体的检测

目的分析内蒙地区发热患者中冠状病毒的感染情况。方法以SARS冠状病毒感染Vero细胞涂片为冠状病毒抗原片,用间接免疫荧光法分别检测55例发热患者和68例正常人血清中冠状病毒的IgG、IgM抗体。结果发热患者血清中冠状病毒IgG抗体和IgM抗体阳性率分别为29.1%(16/55)和10.9%(6/55),而正常人血清中只检测到2.9%(2/68)的IgG抗体,且未检测到IgM抗体,2组患者的IgG和IgM抗体阳性率比较差异均有显著性;随机选取7例患者的IgG阳性血清进行SRAS冠状病毒的特异性抗体封闭实验,结果有6例血清仍为阳性,有1例血清转为阴性,说明冠状病毒IgG抗体阳性血清中85.7%为普通冠状病毒特异性,14.3%为SARS冠状病毒特异性。结论普通冠状病毒是内蒙地区发热患者的主要病原体之一,部分患者还存在SARS冠状病毒的既往感染。     

A biochemical comparison of normal human liver and hepatocellular carcinoma ferritins.

1. The iron contents, gel migration rates and isoelectric-focusing patterns of normal liver and hepatocellular carcinoma ferritins from the same patients were compared. 2. Sucrose-density-gradient centrifugation showed that the number of iron atoms per ferritin molecule was decreased to approximately half in carcinoma tissue when compared with normal liver. 3. On electrophoresis, hepatocellular carcinoma ferritin migrates faster and is therefore more negatively charged than normal liver ferritin, thus refuting the general view that the more negatively charged a ferritin molecule the greater its iron content. 4. Comparison of tumour and normal liver ferritin subunit compositions on acid/urea/polyacrylamide gels showed hepatocellular carcinoma ferritin to contain an additional, more negatively charged, subunit to normal liver ferritin. 5. Isoelectric focusing showed that hepatocellular carcinoma tissue contains isoferritins with isoelectric points intermediate between the ranges of normal liver and normal heart isoferritins.     

Erythrocyte ferritin content in idiopathic haemochromatosis and alcoholic liver disease with iron overload.

The erythrocyte ferritin content was measured in patients with either idiopathic haemochromatosis or alcoholic liver disease and iron overload to define its value as a marker for an excess of tissue iron. The mean erythrocyte ferritin content in patients with untreated idiopathic haemochromatosis was increased 60-fold and fell with phlebotomy. After phlebotomy many patients had an increased red cell ferritin content despite normal serum ferritin concentrations. That this reflected persistent iron overload with inadequate phlebotomy was suggested by the higher serum iron concentrations, percentage transferrin saturation, and urinary excretion of iron after administration of desferrioxamine, together with a lower annual iron loss by phlebotomy in this group compared with patients with treated disease and normal red cell ferritin content. The mean erythrocyte ferritin content in patients with alcoholic liver disease and iron overload was increased only sevenfold, and the ratio of erythrocyte to serum ferritin clearly discriminated these patients from those with idiopathic haemochromatosis. The determination of erythrocyte ferritin content is a useful non-invasive test for diagnosing idiopathic haemochromatosis, monitoring the effect of phlebotomy in this disorder, and distinguishing patients with this disorder from those with alcoholic liver disease with iron overload.     

Purification and Characterization of Canine Serum Ferritin-binding Proteins

Ferritin-binding protein (FBP) is known to interact with circulating ferritins in mammals. Canine FBPs were purified from canine serum by affinity chromatography and were identified as IgM, IgG, and IgA by immunoblotting with alkaline phosphatase-labeled antibodies to canine IgM, IgG, and IgA heavy chains. Following further purification by application to a Sephacryl S-300 column, canine FBPs were separated into 81.3- and 27.7-kDa bands by sodium dodecyl sulfate-polyacryamide gel electrophoresis, and the 81.3-kDa band reacted with the anti-canine IgM heavy chain antibody. Purified canine FBP bound to canine liver ferritin, but not to canine albumin and transferrin. FBP showed greater binding to the expressed bovine ferritin H-chain homopolymer than to the expressed bovine ferritin L-chain homopolymer. The binding of FBP with canine liver ferritin was dose-dependently inhibited by anti-rat liver ferritin antibody, and the anti-ferritin antibody dissociated the bound FBP in a dose-dependent manner, even after binding FBP with liver ferritin. The canine ferritin H subunit peptide fragment with amino acid residues 148–155 (NH₂-GDHVTNLR-COOH) in its C-terminal region was recognized by FBP. These results indicate that canine serum FBPs are autoantibodies to ferritin (IgM, IgG, and IgA) and that anti-ferritin autoantibody (IgM) recognizes the C-terminal region of ferritin H subunit.