Quantifying Aotus monkey cytokines by real-time quantitative RT-PCR

Coaña, Yago Pico de; Barrero, Carlos; Cajiao, Isabela; Mosquera, Catalina; Patarroyo, Manuel Elkín

doi:10.1016/j.cyto.2004.04.004

Cited by 10 publications

(4 citation statements)

References 12 publications

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“…Several commercially available methods have been used for detecting TNFα: enzyme-linked immunosorbent assays (ELISA), , radioimmunoassay (RIA), cytotoxicity assay, flow cytometry, and RT-PCR . The ELISA method offers high sensitivity with a detection limit of 5 pg/mL. , However, these conventional methods typically involve multiple staining and washing steps, which are time-consuming and cannot be used in quantitative analysis of TNFα in real time.…”

Section: Introductionmentioning

confidence: 99%

Photostable Single-Molecule Nanoparticle Optical Biosensors for Real-Time Sensing of Single Cytokine Molecules and Their Binding Reactions

2008

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We synthesized tiny stable silver nanoparticles (2.6 ± 1.1 nm) and used its small surface area and functional groups to control single molecule detection (SMD) volumes on single nanoparticles. These new approaches allowed us to develop intrinsic single molecule nanoparticle optical biosensors (SMNOBS) for sensing and imaging of single human cytokine molecules, recombinant human tumor necrosis factor-α (TNFα), and probing its binding reaction with single monoclonal antibody (MAB) molecules in real-time. We found that SMNOBS retained their biological activity over months and showed exceptionally high photostability. Our study illustrated that smaller nanoparticles exhibited higher dependence of optical properties on surface functional groups, making it a much more sensitive biosensor. Localized surface plasmon resonance spectra (LSPRS) of SMNOBS showed a large red shift of peak wavelength of 29 ± 11 nm, as single TNFα molecules bound with single MAB molecules on single nanoparticles. Utilizing its LSPRS, we quantitatively measured its binding reaction in real time at SM level, showing stochastic binding kinetics of SM reactions with binding equilibrium times ranging from 30 to 120 min. SMNOBS exhibited extraordinarily high sensitivity and selectivity, and a notably wide dynamic range of 0-200 ng/mL (0-11.4 nM). Thus, SMNOBS is well suited for the fundamental study of biological functions of single protein molecules and SM interactions of chemical functional groups with the surface of nanoparticles, as well as development of effective disease diagnosis and therapy.

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Section: Introductionmentioning

confidence: 99%

Photostable Single-Molecule Nanoparticle Optical Biosensors for Real-Time Sensing of Single Cytokine Molecules and Their Binding Reactions

2008

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“…However, conserved HABPs were found to be neither antigenic nor immunogenic or protection inducers when tested in Aotus monkeys, a non-human primate model highly susceptible to human malarias [8], [9] and whose immune system molecules share a high degree of similarity with their human counterparts, specially with those involved in antigen presentation such as α/β TCRs [10], [11] and MHCII–HLA-DRβ1*-like molecules (88% to 100% similarity is reported for the Peptide binding region or PBR of these molecules) [12].…”

Section: Introductionmentioning

confidence: 99%

3D Analysis of the TCR/pMHCII Complex Formation in Monkeys Vaccinated with the First Peptide Inducing Sterilizing Immunity against Human Malaria

et al. 2010

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T-cell receptor gene rearrangements were studied in Aotus monkeys developing high antibody titers and sterilizing immunity against the Plasmodium falciparum malaria parasite upon vaccination with the modified synthetic peptide 24112, which was identified in the Merozoite Surface Protein 2 (MSP-2) and is known to bind to HLA-DRβ1*0403 molecules with high capacity. Spectratyping analysis showed a preferential usage of Vβ12 and Vβ6 TCR gene families in 67% of HLA-DRβ1*0403-like genotyped monkeys. Docking of peptide 24112 into the HLA-DRβ1*0401–HA peptide–HA1.7TCR complex containing the VDJ rearrangements identified in fully protected monkeys showed a different structural signature compared to nonprotected monkeys. These striking results show the exquisite specificity of the TCR/pMHCII complex formation needed for inducing sterilizing immunity and provide important hints for a logical and rational methodology to develop multiepitopic, minimal subunit-based synthetic vaccines against infectious diseases, among them malaria.

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“…IFN-γ, IL-4, IL-10 and IL-13 have close relationship with the immune response. IL-4 and IL-13 have the function of promoting production of IgE [3] [4]. The anatomical structure and the physiological characteristics of macaques are similar to those of humans.…”

Section: Introductionmentioning

confidence: 74%

Establishment of Cytokines Quantitative PCR Methods and Its Application in Anaphylaxis Study of Houttuynia Cordata Injection

Zeng¹,

Sun²,

Cheng³

et al. 2015

JBM

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We design specific primers, according to the four kinds of important macaque cytokines, based on macaque β-action genes, then extract total RNA in peripheral blood mononuclear cells of macaques, and trascriptase cDNA reversely after establishing real-time quantitative PCR, and test the relative expression amount of several kinds of important cytokines during the hypersensitive process. The testing method of quantitative PCR in this experiment is highly sensitive, all cytokines are up to 102 copies/μL and highly reproducible, the intraassay and interassay coefficients of variation are less than 6%. The method applied to the sensitive reaction experiment on cytokines induced by houttuynia cordata turns out that the cytokines relative expression of houttuynia cordata distillate with low dose group and high dose group is significantly higher than that of saline group (P > 0.01). Meanwhile, significantly increased IL-4, IL-10 and IL-13 and decreased IFN-γ are found in both group A and group B after houttuynia injection (P > 0.05).

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Quantifying Aotus monkey cytokines by real-time quantitative RT-PCR

Cited by 10 publications

References 12 publications

Photostable Single-Molecule Nanoparticle Optical Biosensors for Real-Time Sensing of Single Cytokine Molecules and Their Binding Reactions

Photostable Single-Molecule Nanoparticle Optical Biosensors for Real-Time Sensing of Single Cytokine Molecules and Their Binding Reactions

3D Analysis of the TCR/pMHCII Complex Formation in Monkeys Vaccinated with the First Peptide Inducing Sterilizing Immunity against Human Malaria

Establishment of Cytokines Quantitative PCR Methods and Its Application in Anaphylaxis Study of Houttuynia Cordata Injection

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