Enzyme-linked immunosorbent assay for C1q in human serum by use of monoclonal antibodies

A sandwich ELISA system has been developed for the detection of C1q in human serum. It is specific, uses monoclonal antibodies, is sensitive into the nanogram range and is rapidly performed. Therefore, it may be a helpful tool for clinical routine diagnosis, e.g., detecting abnormal C1q levels in patients with rheumatic disorders. Various combinations of poly- and monoclonal antibodies were tested in a sandwich assay. One of these combinations, in particular, resulted in a highly reproducible standard curve: C1q bound to solid-phase polyclonal anti-C1q was detected by the monoclonal antibody 242 G3. In this assay, the C1q concentration in sera of normal individuals was found to be 160 micrograms/ml (mean value of 70 normal human sera). This ELISA detected nanogram levels of C1q and gave results comparable to those obtained by haemolytic C1q titration. One nanogram of C1q corresponded to ca. 2.6 X 10(10) effective C1q molecules. With this technique, selective C1q deficient sera as well as sera from patients with rheumatoid diseases were analysed.     

A novel strategy for targeting CD4+ PPD-reactive T cells against tumour cells using PPD monoclonal antibody heteroconjugates.

We have constructed PPD monoclonal antibody heteroconjugates specific for a tumour-associated antigen of C57BL/6 melanomas or for human complement component C3d fixed de novo to murine fibrosarcoma cells (MC6A). The ability of our heteroconjugates to target CD4+ PPD-reactive T cells against the appropriate tumour targets was then determined in vitro. Heteroconjugate-treated B16-F10 and MC6A tumour targets were both able to present PPD to the specific T cells, resulting in activation and concomitant lymphokine secretion. Secreted lymphokines were then demonstrated to cause significant tumour cytolysis and cytostasis in vitro. Preliminary experiments in vivo suggest that this targeting system may provide the basis for a future immunotherapeutic strategy.     

NCL-5D3: a new monoclonal antibody recognizing low molecular weight cytokeratins effective for immunohistochemistry using fixed paraffin-embedded tissue

Production of a new monoclonal antibody designated NCL-5D3 is described. The antibody recognizes several low molecular weight cytokeratins, in particular cytokeratin Moll number 8 as determined by immunoblotting studies, and is highly effective for immunocytochemistry using routinely processed paraffin-embedded material. Staining is enhanced by prior treatment of the sections with trypsin. Assessment using a wide variety of normal and neoplastic tissue indicates reactivity with all tissues of simple or glandular epithelial origin, and in addition with many squamous carcinomas. Thus the antibody should prove of value in diagnostic histopathology.     

The use of natural interferon alpha conjugated to a monoclonal antibody anti mammary epithelial mucin (Mc5) for the treatment of human breast cancer xenografts

Summary An immunoconjugate composed of natural interferon (nIFN) bound in a noncleavable fashion to a monoclonal antibody (MoAb) recognizing a breast epithelial membrane mucin (Mc5) was used to treat xenografts of a human mammary carcinoma cell line (MCF-7) growing in nude mice. The immunoconjugate (nIFN/Mc5) was administered as 20 intralesional (i.l.) injections to 1 of 2 xenografts in each animal. It was found that nIFN/Mc5 produced a significant enhancement of the growth inhibitory actions of nIFN on the injected tumors. Further enhancement was obtained when nIFN or nIFN together with Mc5 (at a dose 10 times larger than that present in nIFN/Mc5) were added to the immunoconjugate. Biodistribution experiments showed that the uptake of¹²⁵I-nIFN/Mc5 by the tumors was greater and its elimination slower than for¹²⁵I-nIFN alone or conjugated to irrelevant mouse IgG₁. In addition, the immunoconjugate up-regulated the antigenic expression of a breast epithelial membrane mucin by the carcinoma cells, an up-regulation which was not significantly different from that produced by nIFN alone. The contralateral noninjected tumors exposed to systemic levels of the immunoconjugate showed an enhancement of antitumor effects, but to a lesser extent than the injected tumors. These findings suggest that the enhancement of the growth inhibitory action of the immunoconjugate was related to the specific binding of Mc5 which targeted the IFN to the carcinoma cells and impeded its elimination. It is likely that the targeting was favored by the IFN-mediated up-regulation of antigenic expression by the carcinoma cells, thereby producing a cascade of interrelated effects. The results of this study point out the feasibility and potential usefulness of IFN treatment by means of immunoconjugates as well as the worth of pursuing and improving this form of therapy.     

鼠抗人单克隆抗体LH7：2在营养不良型大疱性表皮松解症诊断中的应用

目的 探讨鼠抗人单克隆抗体LH7：2诊断营养不良型大疱性表皮松解症的意义。方法 采用间接免疫荧光法对临床诊断为营养不良型大疱性表皮松解症（DEB）,分别来源于3个不同家系各1名患者,包括2名显性遗传型营养不良型大疱性表皮松解症（dominant DEB,DDEB)和1名隐性遗传型营养不良型大疱性表皮松解症（recessive DEB,RDEB)进行皮肤基底膜带（BMZ）Ⅶ型胶原检测,同时选择单纯型大疱性表皮松解症（EBS）、大疱性类天疱疮及正常人各1名做对照。结果 3名对照BMZ免疫荧光阳性,2名DDEB荧光均减弱,RDEB荧光阴性。结论 鼠抗人单克隆抗体LH7：2诊断营养不良性大疱性表皮松解症有重要意义。     

Chemical conjugation of a novel antibody-interleukin 2 immunoconjugate agains t c-erbB-2 product

Objective To develop a new chemical method to produce a monoclonal antibody (MoAb) 520C9/recombinant human interleukin 2 (rhIL-2) conjugate. Methods MoAb 520C9 reactive with the protooncogene c-erbB-2 product P185 was chemically conjugated with rhIL-2 by using a simple two-step method.First, the rhIL-2 was activated by Sulfosuccinimidyl 4-［N-maleimidomethyl］ cyclohexane-1-carboxylate, a heterobifunctional linker, and N-succinimidyl s-acetylthioacetate was introduced onto 520C9.Then SATA on the 520C9 was reacted with the maleimide group on the activated rhIL-2 to generate 520C9-rhIL-2 immunoconjugate. Results The immunoconjugate retained the antigen binding activity compared to the respective native antibody as determined by an indirect live cell binding assay.The immunoconjugate also possessed IL-2 activity as measured by the standard CTLL-2 cells proliferation assay and the stimulation of human peripheral blood mononuclear cells (PBMCs) into lymphokine-activated killer cells. Conclusion Our method of conjugation of rhIL-2 to 520C9 preserves the binding activity of the antibody and the cytokine function of IL-2.This simple and efficient method of conjugation should be applicable to other types of MoAbs and recombinant cytokines.