The glycoprotein of measles virus. External radioactive labelling of its carbohydrate and partial characterization of the glycopeptide

Measles virus was propagated in VERO cells and purified from the culture supernatants by two successive tartrate-density-gradient centrifugations. Surface carbohydrates were labelled both in vitro and in vivo with 3H after treatment with galactose oxidase/NaB3H4 or with [3H]glucosamine. The major labelled glycoprotein in measles virions had a mol.wt. of 79 000. After labelling with periodate/NaB3H4, which would result in specific labelling of sialic acid residues, the 79 000-mol.wt. glycoprotein was very weakly labelled. This suggests that there is no or a very low amount of sialic acid in the virions. Further analysis of the glycoprotein showed that galactose is the terminal carbohydrate unit in the oligosaccharide, and the molecular weight of the glycopeptide obtained after Pronase digestion is about 3000. The oligosaccharide is attached to the polypeptide through an alkali-stable bond, indicating a N-glycosidic asparagine linkage.

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Measles Virus Attenuation Associated with Transcriptional Impediment and a Few Amino Acid Changes in the Polymerase and Accessory Proteins

Journal of Virology ◽

10.1128/jvi.72.11.8690-8696.1998 ◽

1998 ◽

Vol 72 (11) ◽

pp. 8690-8696 ◽

Cited By ~ 72

Author(s):

Makoto Takeda ◽

Atsushi Kato ◽

Fumio Kobune ◽

Hiroko Sakata ◽

Yan Li ◽

...

Keyword(s):

Amino Acid ◽

Vero Cell ◽

Cell Line ◽

Measles Virus ◽

Vero Cells ◽

Functional Difference ◽

Pathogenic Potential ◽

P Proteins

ABSTRACT Measles virus (MV) isolated in B95a cells, a marmoset B-cell line, retains full pathogenicity for cynomolgus monkeys, while its derivative obtained by adaptation to the growth in Vero cells, a monkey kidney cell line, loses the pathogenic potential (F. Kobune, H. Sakata, and A. Sugiura, J. Virol. 64:700–705, 1990). Here, we show with a pair of strains, a fresh isolate (9301B) in B95a cells and its Vero cell-adapted form (9301V), that the in vivo attenuation parallels the decrease of replication and syncytium-inducing capabilities in the original B95a cells and that these in vitro phenotypes are attributable to impediment of transcription, which is already obvious at the level of primary transcription catalyzed by the virion-associated RNA polymerase. On the other hand, cell fusion assays detected no functional difference between the glycoproteins of the two viruses. Essentially the same transcriptional impediment with reduced syncytium induction following Vero cell adaptation was found with two other pairs of strains that had been similarly prepared. Nucleotide sequence comparison between the 9301B and 9301V viruses revealed that a few (at most five) amino acid changes, which sporadically took place in the polymerase (L and P proteins) and/or accessory V and C proteins, were responsible for the in vitro and in vivo attenuation through adaptation to growth in Vero cells.

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432. Characterization of a Novel Oncolytic Measles Virus Retargeted Against the Urokinase Receptor: In Vitro and In Vivo Studies

Molecular Therapy ◽

10.1016/j.ymthe.2006.08.498 ◽

2006 ◽

Vol 13 ◽

pp. S166-S167

Author(s):

Jaime R. Merchan ◽

Caili Tong ◽

Takafumi Nakamura ◽

Ianko Iankov ◽

Stephen J. Russell

Keyword(s):

Measles Virus ◽

Urokinase Receptor ◽

In Vivo Studies

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Composition and Clinical Significance of Exosomes in Tuberculosis: A Systematic Literature Review

Journal of Clinical Medicine ◽

10.3390/jcm10010145 ◽

2021 ◽

Vol 10 (1) ◽

pp. 145

Author(s):

Fantahun Biadglegne ◽

Brigitte König ◽

Arne C. Rodloff ◽

Anca Dorhoi ◽

Ulrich Sack

Keyword(s):

Vaccine Development ◽

Biological Fluids ◽

Diagnostic Tools ◽

Major Health ◽

Fast Detection ◽

Isolation And Characterization ◽

Culture Supernatants

Tuberculosis (TB) remains a major health issue worldwide. In order to contain TB infections, improved vaccines as well as accurate and reliable diagnostic tools are desirable. Exosomes are employed for the diagnosis of various diseases. At present, research on exosomes in TB is still at the preliminary stage. Recent studies have described isolation and characterization of Mycobacterium tuberculosis (Mtb) derived exosomes in vivo and in vitro. Mtb-derived exosomes (Mtbexo) may be critical for TB pathogenesis by delivering mycobacterial-derived components to the recipient cells. Proteomic and transcriptomic analysis of Mtbexo have revealed a variety of proteins and miRNA, which are utilized by the TB bacteria for pathogenesis. Exosomes have been isolated in body fluids, are amenable for fast detection, and could contribute as diagnostic or prognostic biomarker to disease control. Extraction of exosomes from biological fluids is essential for the exosome research and requires careful standardization for TB. In this review, we summarized the different studies on Mtbexo molecules, including protein and miRNA and the methods used to detect exosomes in biological fluids and cell culture supernatants. Thus, the detection of Mtbexo molecules in biological fluids may have a potential to expedite the diagnosis of TB infection. Moreover, the analysis of Mtbexo may generate new aspects in vaccine development.

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Characterization of Measles Virus-Specific Proteins Synthesized In Vivo and In Vitro from Acutely and Persistently Infected Cells

Journal of Virology ◽

10.1128/jvi.29.3.1099-1106.1979 ◽

1979 ◽

Vol 29 (3) ◽

pp. 1099-1106 ◽

Cited By ~ 10

Author(s):

Shmuel Rozenblatt ◽

Marian Gorecki ◽

Helen Shure ◽

Carol L. Prives

Keyword(s):

Measles Virus ◽

Persistently Infected ◽

Specific Proteins ◽

Infected Cells

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Virulence parameters in the characterization of strains of Entamoeba histolytica

Revista do Instituto de Medicina Tropical de São Paulo ◽

10.1590/s0036-46651997000200001 ◽

1997 ◽

Vol 39 (2) ◽

pp. 65-70 ◽

Cited By ~ 2

Author(s):

Maria A GOMES ◽

Maria N. MELO ◽

Gil P.M. PENA ◽

Edward F. SILVA

Keyword(s):

Entamoeba Histolytica ◽

Cytopathic Effect ◽

Axenic Culture ◽

Vero Cells ◽

In Vitro Assays ◽

Biological Assays ◽

Restriction Fragment Pattern

Differences in virulence of strains of Entamoeba histolytica have long been detected by various experimental assays, both in vivo and in vitro. Discrepancies in the strains characterization have been arisen when different biological assays are compared. In order to evaluate different parameters of virulence in the strains characterization, five strains of E. histolytica, kept under axenic culture, were characterized in respect to their, capability to induce hamster liver abscess, erythrophagocytosis rate and cytopathic effect upon VERO cells. It was found significant correlation between in vitro biological assays, but not between in vivo and in vitro assays. Good correlation was found between cytopathic effect and the mean number of uptaken erythrocytes, but not with percentage of phagocytic amoebae, showing that great variability can be observed in the same assay, according to the variable chosen. It was not possible to correlate isoenzyme and restriction fragment pattern with virulence indexes since all studied strains presented pathogenic patterns. The discordant results observed in different virulence assays suggests that virulence itself may not the directly assessed. What is in fact assessed are different biological characteristics or functions of the parasite more than virulence itself. These characteristics or functions may be related or not with pathogenic mechanisms occurring in the development of invasive amoebic disease

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Influence of Platelet Membrane Sialic Acid and Platelet-Associated IgG on Ageing and Sequestration of Blood Platelets in Baboons

Thrombosis and Haemostasis ◽

10.1055/s-0038-1649648 ◽

1993 ◽

Vol 70 (04) ◽

pp. 676-680 ◽

Cited By ~ 24

Author(s):

H F Kotzé ◽

V van Wyk ◽

P N Badenhorst ◽

A du P Heyns ◽

J P Roodt ◽

...

Keyword(s):

Sialic Acid ◽

Life Span ◽

Blood Platelets ◽

Senescent Cell ◽

Platelet Membrane ◽

Antigen Complex ◽

The Mean ◽

Aggregation Of Platelets

SummaryPlatelets were isolated from blood of baboons and treated with neuraminidase to remove platelet membrane sialic acid, a process which artificially ages the platelets. The platelets were then labelled with 111In and their mean life span, in vivo distribution and sites of Sequestration were measured. The effect of removal of sialic acid on the attachment of immunoglobulin to platelets were investigated and related to the Sequestration of the platelets by the spleen, liver, and bone marrow. Removal of sialic acid by neuraminidase did not affect the aggregation of platelets by agonists in vitro, nor their sites of Sequestration. The removal of 0.51 (median, range 0.01 to 2.10) nmol sialic acid/108 platelets shortened their life span by 75 h (median, range 0 to 132) h (n = 19, p <0.001), and there was an exponential correlation between the shortening of the mean platelet life span and the amount of sialic acid removed. The increase in platelet-associated IgG was 0.112 (median, range 0.007 to 0.309) fg/platelet (n = 25, p <0.001) after 0.79 (median, range 0.00 to 6.70) nmol sialic acid/108 platelets was removed (p <0.001). There was an exponential correlation between the shortening of mean platelet life span after the removal of sialic acid and the increase in platelet-associated IgG. The results suggest that platelet membrane sialic acid influences ageing of circulating platelets, and that the loss of sialic acid may have exposed a senescent cell antigen that binds IgG on the platelet membrane. The antibody-antigen complex may then provide a signal to the macrophages that the platelet is old, and can be phagocytosed and destroyed.

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In vivo and in vitro characterization of immediate release dosage forms containing n-acetylcysteine using the dynamic open flow-through test apparatus

10.26226/morressier.57d6b2b7d462b8028d88dd29 ◽

2016 ◽

Author(s):

Maximilian Sager

Keyword(s):

Immediate Release ◽

Dosage Forms ◽

Test Apparatus ◽

Open Flow ◽

In Vitro Characterization ◽

Flow Through

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Rapid, Refined, and Robust Method for Expression, Purification, and Characterization of Recombinant Human Amyloid-beta M1-42

10.26434/chemrxiv.7725002.v1 ◽

2019 ◽

Author(s):

Priya Prakash ◽

Travis Lantz ◽

Krupal P. Jethava ◽

Gaurav Chopra

Keyword(s):

Amyloid Beta ◽

Western Blots ◽

Force Microscopy ◽

E Coli ◽

Atomic Force ◽

Set Up ◽

Short Time

Amyloid plaques found in the brains of Alzheimer’s disease (AD) patients primarily consists of amyloid beta 1-42 (Ab42). Commercially, Ab42 is synthetized using peptide synthesizers. We describe a robust methodology for expression of recombinant human Ab(M1-42) in Rosetta(DE3)pLysS and BL21(DE3)pLysS competent E. coli with refined and rapid analytical purification techniques. The peptide is isolated and purified from the transformed cells using an optimized set-up for reverse-phase HPLC protocol, using commonly available C18 columns, yielding high amounts of peptide (~15-20 mg per 1 L culture) in a short time. The recombinant Ab(M1-42) forms characteristic aggregates similar to synthetic Ab42 aggregates as verified by western blots and atomic force microscopy to warrant future biological use. Our rapid, refined, and robust technique to purify human Ab(M1-42) can be used to synthesize chemical probes for several downstream in vitro and in vivo assays to facilitate AD research.

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