Partial molecular cloning of the JHK retrovirus using gammaretrovirus consensus PCR primers.

The JHK virus (JHKV) was previously described as a type C retrovirus that has some distinctive ultrastructural features and replicates constitutively in a human B-lymphoblastoid cell line, JHK-3. In order to facilitate the cloning of sequences from JHKV, a series of partially degenerate consensus retroviral PCR primers were created by a data-driven design approach based on an alignment of 14 diverse gammaretroviral genomes. These primers were used in the PCR amplification of purified JHK virion cDNA, and ana lysis of the resulting amplified sequence indicates that the JHKV is in the murine leukemia virus (MLV) family.

Frequency and magnitude of interferon β neutralizing antibodies in the evaluation of interferon β immunogenicity in patients with multiple sclerosis.

Patients with multiple sclerosis (MS) treated with interferon β (IFNβ) preparations develop varying levels of antibodies that neutralize the biological effects of IFNβ, reduce its in vivo bioavailability, and diminish its therapeutic efficacy. The aim was to determine as distinct measures of immunogenicity the occurrence (frequency) and the magnitude (level) of IFNβ neutralizing antibody (NAb) formation in a large Canadian population as a cross-sectional study of patients with MS treated in a clinical practice setting with different, equally available IFNβ products: Avonex(®) (intramuscular IFNβ-1a), Rebif(®) (subcutaneous (SC) IFNβ-1a) at 22 and 44 μg, and Betaseron(®) (SC IFNβ-1b). Over a 3-year period 3,124 serum samples from 2,711 patients with MS were submitted by neurologists in MS clinics distributed across Canada and tested for NAbs in a single independent laboratory, utilizing a quantitative, standardized NAb bioassay.

Quantification of the neutralization of cytokine biological activity by antibody: the ten-fold reduction bioassay of interleukin-6 as growth factor.

The measurement of neutralizing antibodies (NAbs) to biological therapeutic agents is important clinically as well as for the preclinical evaluation of product immunogenicity. To determine whether the theoretical concepts and experimental data from studies of the nature of antibody neutralization of interferons (IFNs) can apply to unrelated protein effector molecules, neutralization experiments were undertaken with interleukin-6 (IL-6), a proinflammatory, highly pleiotropic cytokine. By following IL-6 induction of hybridoma cell growth, we demonstrated that anti-IL-6 monoclonal and polyclonal NAbs can be measured with a bioassay design structured to reduce 10 Laboratory Units (LU)/mL to 1 LU/mL.

The neutralization of interferons by antibody III. The constant antibody bioassay, a highly sensitive quantitative detector of low antibody levels.

The neutralizing antibodies (NAbs) that develop in patients during interferon (IFN) therapy can reduce its beneficial effects. The universally employed method of NAb measurement currently is the constant IFN method, in which antigen at a single given concentration is mixed with serial dilutions of serum, the lowest final dilution of which (usually 1:20) is constrained by the potential adverse effect of human serum on human cells in culture. The constant antibody (Ab) method described herein uses serum at a certain set dilution (usually 1:20) mixed with a series of IFN concentrations.

Validating parameters of a luciferase reporter gene assay to measure neutralizing antibodies to IFNbeta in multiple sclerosis patients.

Neutralizing antibodies (NAbs) can occur in some multiple sclerosis (MS) patients receiving interferon beta (IFNbeta) therapy. NAbs reduce drug bioavailabity and high NAb titers reduce drug efficacy. We describe the validation of the R.

Urea-nuclease treatment of concentrated retrovirions preserves viral RNA and removes polymerase chain reaction-amplifiable cellular RNA and DNA.

Cellular nucleic acids can interfere with the molecular cloning of retroviruses, a problem that is particularly serious with viruses propagated in lymphoblastoid cells that release large amounts of microvesicles and other cellular components. The approach taken to circumvent such problems involved first suspending viral pellets in water to allow any residual microvesicles to swell and perhaps lyse during overnight or longer incubation periods. Urea was then added to a concentration of 1.

How the American Society for Virology was founded.

The American Society for Virology, the very first such Society to be formed anywhere, was founded at a meeting of some 40 virologists at Chicago O'Hare International airport on June 9, 1981. They met after a decade and a half of intense discussion that originated at the 9th International Congress of Microbiology in Moscow in 1966 when a small group of virologists requested the International Association of Microbiological Societies to form a Virology Section within IAMS, and this request was rejected. Virologists therefore held their own First International Congress of Virology in Helsinki in 1968 which was very successful and generated intense informal discussion among leading virologists in this country as to the desirability of founding an American society for virologists.

Serum neopterin, an immune activation marker, independently predicts disease progression in advanced HIV-1 infection.

Background: CD4+ T lymphocyte (CD4) counts and plasma human immunodeficiency virus (HIV) type 1 RNA concentrations predict clinical outcome in HIV-1 infection. Our objective was to assess the independent prognostic value for disease progression of soluble markers of immune system activation.

Methods: This retrospective marker-validation study utilized previously obtained clinical and laboratory data, including CD4+ cell counts, and made use of stored frozen serum samples to assay for levels of beta2-microglobulin, neopterin, endogenous interferon, triglycerides, interleukin-6, soluble tumor necrosis factor- alpha receptor II, and HIV-1 RNA, and to determine HIV genotypic reverse-transcriptase inhibitor resistance.

Perspectives on the neutralization of interferons by antibody.

Interferon (IFN) neutralizing antibody (NAb) interferes with the binding of the IFN molecule to its cellular receptor, thereby preventing receptor activation required to trigger the cascade of signal transduction events that lead to the varied actions of IFN. IFN antibody neutralization is measured in an IFN bioassay using cultured human cells. Antibody results need to be represented in a reproducibly quantitative manner to know what proportion of patients treated with an IFN preparation form NAbs and whether the levels of antibody measured are likely to have clinically adverse effects; however, antibody titers have been reported in various ways.

Neutralization of the biological activity of cytokines and other protein effectors by antibody: theoretical formulation of antibody titration curves in relation to antibody affinity.

Patients treated with interferons, other cytokines, or various biologically active proteins may form neutralizing antibodies, which can adversely affect clinical outcome. It is therefore important to understand how antibodies neutralize such soluble protein antigens and how best to quantitate such antibodies. By applying the mass action law to antigen-antibody reactions, we previously developed a mathematical model applicable in two situations: first, for antibodies having low affinity for the antigen concerned (the Constant Proportion (CP) case), and, second, for antibodies having high affinity (the Fixed Amount (FA) case).

Molecular cloning, complete sequence, and biological characterization of a xenotropic murine leukemia virus constitutively released from the human B-lymphoblastoid cell line DG-75.

A previously undetected retrovirus has been isolated from the human Epstein-Barr virus (EBV)-negative, B-lymphoblastoid DG-75 cell line, widely used for EBV gene transfection studies. The complete 8207-base genome of the DG-75 retrovirus was molecularly cloned from viral mRNA and sequenced (Accession No. AF221065).