新城疫病毒单克隆抗体的特性及应用

建立了8个分泌抗新城疫病毒(NDV)特异性单克隆抗体(McAb)的杂交瘤细胞株,根据它们的免疫生物学特性可以分为三种类型:(1)具有FA和ELISA特性(FN1、FN4、FN29、FN30、FN35、FNl22);(2)具有FA、ELISA和HI特性(FN7);(3)具有ELISA、HI特性和中和能力(FN106),根据FN30和FN106的ELISA试验,可将11个NDV毒株分为二种不同的抗原群,应用FN4-FITC,FN7-FITC和FN29-HRP试剂,对人工感染NDV和野外送检病例检测结果表明,单抗试剂的DFA阳性率(92.3％)高于病毒分离阳性率(87.2％),两种方法的符合率89.7％,这些单抗试剂用于临床诊断敏感性和特异性高,且方法快速、简便。     

广谱肾综合征出血热病毒单克隆抗体的A35的生物学性状

具有中和及血凝抑制活性的、能和世界各地分离到的肾综合征出血热病毒(HFRSV)发生反应的、广谱的单克隆抗体(McAb),对HFRSV的诊断和分子生物学研究都有重要意义。 本文着重比较了HFRSV McAbA5、A19、A25-1、A25-7和A35的生物学性状,并观察了对感染动物的实验治疗效果。     

Inhibition of in vitro fertilization by mouse anti-mouse sperm sera and preliminary antigen identification

Antisperm antibodies are implicated as one causative factor of infertility, but the target antigens have not been identified. Immune responses to sperm antigens are qualitatively variable even within a single mouse strain. We took advantage of this variability and immunized individual female mice to allogeneic sperm to reflect their natural exposure during mating. We determined the ability of the individual sera to inhibit in vitro fertilization and to bind to sperm antigens separated by electrophoresis. Compared to preimmune sera, four of five immune sera significantly inhibited in vitro fertilization. The serum from individual mice bound variable panels of sperm antigens. By comparing the panels, we identified two polypeptides with molecular weights of 40,000 and 44,000 that were bound by all sera. We propose that these molecules may be good candidates for further investigation of the immunoprophylaxis of pregnancy.     

Monoclonal antibodies to rabbit liver cathepsin B

Three stable hybridoma cell lines (AF8, BC11, CE2) have been produced that secrete antibodies specific for cathepsin B. These have been characterized by ELISA, SDS-PAGE immunostaining, immunoprecipitation and immunofluorescent staining. CE2 immunoprecipitated native cathepsin B with retention of enzymic activity, but failed to cross-react with the alkali-denatured enzyme. BC11 bound only to the denatured form of cathepsin B and AF8 cross-reacted with both native and denatured cathepsin B. However, unlike CE2-immunoprecipitated enzyme, activity could be detected only after dissociation of the antigen-AF8 antibody complex. No cross reaction was found with any lysosomal protein includihg the cysteine proteinases, catbepsins H and L.Abbreviations ELISA Enzyme-linked immunoadsorbent assay - EIP Enzyme immunoprecipitation - PAGE polyacrylamide gel electrophoresis - Ep-475 L-trans-epoxysuccinyl-leucylamido (-methyl) butane - Z benzyloxycarbonyl - NMec N-methylcoumarin - PEB phosphate-EDTA-Brij 35 - IAA iodoacetic acid - PBS phosphate-buffered saline - DMEM Dulbecco's Minimal Essential Medium - FITC fluorescein isothiocyanate     

Immunocytochemistry of magnocellular neurons of supraoptic and paraventricular nuclei of normal and Brattleboro rats with vasopressin anti-idiotype antibody

Summary A vasopressin anti-idiotype antibody was generated by immunization with purified IgG of a primary vasopressin antiserum. The anti-idiotype antibody immunostained neurons in the supraoptic and paraventricular nuclei of the hypothalamus of normal and Brattleboro rats. The distribution of immunostained perikarya in these hypothalamic nuclei together with the staining of fibers in median eminence and neural lobe was similar to that observed in normal rats with anti-vasopressin and suggests strongly that vasopressinergic neurons are being stained. Absorption studies with vasopressin and a vasopressin-binding receptor protein further indicate that a receptor associated with vasopressinergic neurons is recognized by the anti-idiotype antibody.Supported by NIH grants ES03239, NS18626 and NSF grant BNS-8310914. D.T.P. is the receipient of RCDA award NS00869     

Morphological evidence for the existence of a more complex cytoskeleton in Amoeba proteus

Summary Various stabilization and extraction procedures were tested to demonstrate the ultrastructural organization of the cytoskeleton in normal, locomoting Amoeba proteus. Most reliable results were obtained after careful fixation in glutaraldehyde/lysine followed by prolonged extraction in a polyethylene glycol/Triton X-100 solution. Before dehydration in a graded series of ethanol and critical-point drying, the amoebae were split by the sandwich-technique, i.e., by mechanical cleavage of cells mounted between two poly-L-lysine-coated glass slides. Platinum-carbon replicas as well as thin sections prepared from such cell fragments revealed a cytoskeleton composed of at least four different types of filaments: (1) 5–7-nm filaments organized as a more or less ordered cortical network at the internal face of the plasma membrane and probably representing F-actin; (2) 10–12-nm filaments running separately or slightly aggregated through the cytoplasm and probably representing intermediate filaments; (3) 24–26-nm filaments forming a loose network and probably representing microtubules; and (4) 2–4-nm filaments as connecting elements between the other cytoskeleton constituents. Whereas microfilaments are responsible for protoplasmic streaming and other motile phenomena, the function of intermediate filaments and cytoplasmic microtubules in amoebae is still obscure.     

Isolation of low-copy-number sequences that neighbor satellite DNA in mammals

To investigate the role of satellite DNA in eukaryotic genomes, we isolated from an African green monkey (Cercopithecus aethiops) genomic library cloned segments containing the previously described deca-satellite linked to low-copy-number genomic sequences. Three such clones were obtained. The low-copy-number sequences in the three clones do not cross-hybridize suggesting that they derive from different genomic loci. The structure of one of the clones, λAMkA, is described in detail. Subcloned segments containing the low-copy-number sequences from λAMkA anneal to monkey, human and mouse genomic DNA. The subcloned probes were used to select clones containing homologous sequences from a second, independent monkey library as well as from human and mouse genomic libraries. Several of the newly isolated monkey clones hybridized to probes containing the species-specific deca- and -satellites, confirming the genomic association of the low-copy-number sequence in λAMkA with satellite DNA. Moreover, several of the human and mouse clones hybridized to species-specific human and mouse satellite DNAs, respectively. These experiments indicate that the low-copy-number sequence in λMkA and its association with satellite DNA is conserved in primates and rodents.     

Recovery of proteins on a milligram scale from polyacrylamide electrophoresis gels,exemplified by purification of a retinol-binding protein

An apparatus suitable for the recovery of proteins from polyacrylamide gels on a milligram scale by displacement electrophoresis (isotachophoresis) is described along with a buffer system that is suitable for this purpose with most proteins. The technique is illustrated by the recovery of a protein from a 15% polyacrylamide gel. The recovery was almost quantitative and the eluted protein showed little contamination upon quantitative amino acid analysis and automatic Edman degradation.     

Monoclonal antibodies modify acetylcholine-induced ionic channel properties in cultured chick myoballs

Summary Monoclonal antibodies directed against the cholinergic binding site of the acetylcholine receptor were found to alter the ion channel properties in cultured chick myoballs. Time and dose dependent reduction in acetylcholine sensitivity was observed. Noise analysis experiments indicated a decrease in the mean single channel conductance and an increase in the mean single channel open time.     

Isoelectric focusing in immobilized pH gradients: generation and optimization of wide pH intervals with two-chamber mixers

A new technique for generating extended pH gradients (5 pH units) in Immobiline gels is reported. The previously described (J. Biochem. Biophys. Methods 7, 1983, 123-142) five-chamber gradient mixer has been replaced by a two-vessel device. A single mixture of the available Immobilines (pK 3.6, 4.6, 6.2, 7.0, 8.5 and 9.3) is made, with relative concentrations adjusted so as to produce the most uniform buffering power throughout the desired pH interval. This mixture is then divided into two portions, which are titrated to the extremes of the required pH span with an acidic titrant (Immobiline pK approximately 1) and a basic species (Immobiline pK 9.95). Highly reproducible pH gradients (pH 4-9) are thus generated, which appear extremely useful for the first dimensioned of 2-dimensional techniques. Our previously reported computer program has been implemented with an optimization algorithm which, given any cocktail of Immobilines, automatically adjusts the relative initial concentrations until the smoothest possible beta power is found. For the first time it is possible to perform IEF under controlled physico-chemical parameters: pH span and linearity, beta power, ionic strength and molarity of the buffering species.