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International Journal of Nanomedicine 2020Nanocarriers could deliver significantly higher amounts of antigen to antigen-presenting cells (APCs), which have great potential to stimulate humoral and cellular...
BACKGROUND
Nanocarriers could deliver significantly higher amounts of antigen to antigen-presenting cells (APCs), which have great potential to stimulate humoral and cellular response in cancer immunotherapy. Thereafter, silica solid nanosphere (SiO) was prepared, and a model antigen (ovalbumin, OVA) was covalently conjugated on the surface of SiO to form nanovaccine (OVA@SiO). And the application of OVA@SiO for cancer immunotherapy was evaluated.
MATERIALS AND METHODS
SiO solid nanosphere was prepared by the Stöber method, then successively aminated by aminopropyltriethoxysilane and activated with glutaraldehyde. OVA was covalently conjugated on the surface of activated SiO to obtain nanovaccine (OVA@SiO). Dynamic light scattering, scanning electron microscope, and transmission electron microscope were conducted to identify the size distribution, zeta potential and morphology of OVA@SiO. The OVA loading capacity was investigated by varying glutaraldehyde concentration. The biocompatibility of OVA@SiO to DC2.4 and RAW246.7 cells was evaluated by a Cell Counting Kit-8 assay. The uptake of OVA@SiO by DC2.4 and its internalization pathway were evaluated in the absence or presence of different inhibitors. The activation and maturation of bone marrow-derived DC cells by OVA@SiO were also investigated. Finally, the in vivo transport of OVA@SiO and its toxicity to organs were appraised.
RESULTS
All results indicated the successful covalent conjugation of OVA on the surface of SiO. The as-prepared OVA@SiO possessed high antigen loading capacity, which had good biocompatibility to APCs and major organs. Besides, OVA@SiO facilitated antigen uptake by DC2.4 cells and its cytosolic release. Noteworthily, OVA@SiO significantly promoted the maturation of dendritic cells and up-regulation of cytokine secretion by co-administration of adjuvant CpG-ODN.
CONCLUSION
The as-prepared SiO shows promising potential for use as an antigen delivery carrier.
Topics: Adjuvants, Immunologic; Animals; Antigen Presentation; Antigens; Cancer Vaccines; Dendritic Cells; Drug Carriers; Female; Immunotherapy; Mice; Mice, Inbred C57BL; Nanospheres; Oligodeoxyribonucleotides; Ovalbumin; RAW 264.7 Cells; Silicon Dioxide
PubMed: 32368049
DOI: 10.2147/IJN.S242463 -
Bioscience, Biotechnology, and... Aug 1999The recombinant ovalbumin produced in Escherichia coli was purified from the cytoplasmic fraction and analyzed for its chemical and conformational properties. The...
The recombinant ovalbumin produced in Escherichia coli was purified from the cytoplasmic fraction and analyzed for its chemical and conformational properties. The recombinant ovalbumin displayed almost exactly the same circular dichroism and intrinsic tryptophan fluorescence spectra as egg white ovalbumin. As in the egg white protein, four cysteine sulfhydryls and one cystine disulfide were contained in the recombinant protein, according to the results of amino acid analyses; the disulfide bond was found by a peptide mapping analysis to correspond to the native cystine, Cys73-Cys120. According to a gel electrophoresis analysis, the presence of the disulfide bond was accounted for by specific oxidation of the corresponding cysteine residues during purification of the cytoplasmic protein. Unlike the identity in the conformational and peptide structures, none of the post-translational modifications (N-terminal acetylation, phosphorylation, and glycosylation) that are known with egg white ovalbumin were detected in the recombinant protein. The recombinant ovalbumin was transformed into a thermostabilized form in a similar manner to the transformation of egg white protein into S-ovalbumin; alkaline treatment increased the temperature for thermostability by 8.7 degrees C. These data strongly suggest that the post-translational modifications of ovalbumin are not related to the formation mechanism for S-ovalbumin.
Topics: Alkalies; Circular Dichroism; Cytoplasm; Disulfides; Ovalbumin; Peptide Mapping; Protein Conformation; Protein Processing, Post-Translational; Recombinant Proteins; Spectrometry, Fluorescence; Temperature
PubMed: 10501000
DOI: 10.1271/bbb.63.1392 -
Scientific Reports Jun 2020Plant viruses are biologically safe for mammals and can be successfully used as a carrier/platform to present foreign epitopes in the course of creating novel putative...
Plant viruses are biologically safe for mammals and can be successfully used as a carrier/platform to present foreign epitopes in the course of creating novel putative vaccines. However, there is mounting evidence that plant viruses, their virus-like and structurally modified particles may also have an immunopotentiating effect on antigens not bound with their surface covalently. Here, we present data on the adjuvant properties of plant viruses with various shapes (Tobacco mosaic virus, TMV; Potato virus X, PVX; Cauliflower mosaic virus, CaMV; Bean mild mosaic virus, BMMV) and structurally modified TMV spherical particles (SPs). We have analysed the effectiveness of immune response to individual model antigens (ovalbumin, OVA/hen egg lysozyme, HEL) and to OVA/HEL in compositions with plant viruses/SPs, and have shown that CaMV, TMV and SPs can effectively induce total IgG titers to model antigen. Some intriguing data were obtained when analysing the immune response to the plant viruses/SPs themselves. Strong immunity was induced to CaMV, BMMV and PVX, whereas TMV and SPs stimulated considerably lower self-IgG titers. Our results provide new insights into the immunopotentiating properties of plant viruses and can be useful in devising adjuvants based on plant viruses.
Topics: Adjuvants, Immunologic; Animals; Epitopes; Immunization; Mice; Muramidase; Ovalbumin; Plant Viruses
PubMed: 32587281
DOI: 10.1038/s41598-020-67023-4 -
Biosensors Jun 2023Food allergies are an exceptional response of the immune system caused by the ingestion of specific foods. The main foods responsible for allergic reactions are milk,...
Food allergies are an exceptional response of the immune system caused by the ingestion of specific foods. The main foods responsible for allergic reactions are milk, eggs, seafood, soy, peanuts, tree nuts, wheat, and their derived products. Chicken egg ovalbumin (OVA), a common allergen molecule, is often used for the clarification process of wine. Traces of OVA remain in the wine during the fining process, and they can cause significant allergic reactions in sensitive consumers. Consequently, the European Food Safety Authority (EFSA) and the American Food and Drug Administration (FDA) have shown the risks for allergic people to assume allergenic foods and food ingredients, including eggs. Commonly, OVA detection requires sophisticated and time-consuming analytical techniques. Intending to develop a faster assay, we designed a proof-of-concept non-Faradaic impedimetric immunosensor for monitoring the presence of OVA in wine. Polyclonal antibodies anti-OVA were covalently immobilised onto an 11-mercaptoundecanoic-acid (11-MUA)-modified gold surface. The developed immunosensor was able to detect OVA in diluted white wine without the need for an external probe or any pre-treatment step with a sensitivity of 0.20 µg/mL, complying with the limit established by the resolution OIV/COMEX 502-2012 for the quantification of allergens in wine.
Topics: Humans; Ovalbumin; Biosensing Techniques; Wine; Electric Impedance; Immunoassay; Allergens; Food Hypersensitivity
PubMed: 37504068
DOI: 10.3390/bios13070669 -
Frontiers in Immunology 2021CpG-oligodeoxynucleotides (CpG-ODNs) constitute an attractive alternative for asthma treatment. However, very little evidence is available from studies on the oral...
CpG-oligodeoxynucleotides (CpG-ODNs) constitute an attractive alternative for asthma treatment. However, very little evidence is available from studies on the oral administration of CpG-ODNs in animals. Previously, we developed acid-resistant particles (named ODNcap) as an oral delivery device for ODNs. Here, we showed that free feeding of an ODNcap-containing feed prophylactically attenuates allergic airway inflammation, hyperresponsiveness, and goblet cell hyperplasia in an ovalbumin-induced asthma model. Using transcriptomics-driven approaches, we demonstrated that injury of pulmonary vein cardiomyocytes accompanies allergen inhalation challenge, but is inhibited by ODNcap feeding. We also showed the participation of an airway antimicrobial peptide (Reg3γ) and fecal microbiota in the ODNcap-mediated effects. Collectively, our findings suggest that daily oral ingestion of ODNcap may provide preventive effects on allergic bronchopulmonary insults regulation of mechanisms involved in the gut-lung connection.
Topics: Administration, Oral; Animals; Antimicrobial Peptides; Bronchial Hyperreactivity; Female; Gastrointestinal Microbiome; Hypersensitivity; Mice; Mice, Inbred BALB C; Oligodeoxyribonucleotides; Ovalbumin; Pancreatitis-Associated Proteins; Pneumonia
PubMed: 34867960
DOI: 10.3389/fimmu.2021.738041 -
Proceedings of the National Academy of... Dec 1978By recombinant DNA methods, the chicken ovalbumin structural gene has been fused to Escherichia coli lac transcriptional and translational control regions. When a...
By recombinant DNA methods, the chicken ovalbumin structural gene has been fused to Escherichia coli lac transcriptional and translational control regions. When a plasmid containing the hybrid gene was introduced into E. coli, a protein identified as ovalbumin by immunoreactivity and sodium dodecyl sulfate/polyacrylamide gel electrophoresis was synthesized. The chicken ovalbumin made in bacteria was full length (43,000 daltons) and constituted 1.5% of the cellular protein. In addition, the microbially synthesized ovalbumin was secreted through the cell membrane into the periplasmic space of E. coli. The ability of the E. coli secretory apparatus to recognize chicken ovalbumin, which is normally synthesized and secreted in hen oviducts, suggests that common features exist in the secretion-recognition mechanisms found in these two organisms. The bacterial synthesis of significant amounts of chicken ovalbumin demonstrates that the E. coli cellular machinery may be utilized to synthesize a higher eukaryotic protein which is relatively stable in the bacterial intracellular environment.
Topics: Alkaline Phosphatase; Amino Acid Sequence; DNA, Recombinant; Escherichia coli; Lac Operon; Ovalbumin; Plasmids; beta-Galactosidase
PubMed: 104296
DOI: 10.1073/pnas.75.12.5936 -
PloS One 2019The skin is a very suitable organ for the induction of immune responses to vaccine antigens. Antigen delivery systems to the skin by needle and syringe directly deposit...
The skin is a very suitable organ for the induction of immune responses to vaccine antigens. Antigen delivery systems to the skin by needle and syringe directly deposit the antigen into the epidermal-dermal compartment, one of the most immunocompetent sites due to the presence of professional antigen-presenting cells aimed at the induction of antigen-specific T cells. In this study, we analyzed the amount of ovalbumin as an antigen delivered to the skin by a microneedle. When ovalbumin protein as an antigen was delivered to the skin of mice using a dissolving microneedle, it induced an immune response through the enhanced proliferation and cytokines production by the splenocytes and lymph nodes. Also, it effectively increased the ovalbumin-specific CD8+ T cell and CD4+ T cell population and induced an ovalbumin-specific CTL response against the graft of ovalbumin-expressing EG7 tumor cells in the immunized mice. Also, we identified the inhibition of tumor growth and prevention of tumor formation in the context of the therapeutic and prophylactic vaccine, respectively through EG-7 tumor mouse model. Finally, these data show the potential of patches as attractive antigen delivery vehicles.
Topics: Administration, Cutaneous; Animals; Antigens; CD4-Positive T-Lymphocytes; CD8-Positive T-Lymphocytes; Cell Proliferation; Drug Delivery Systems; Immunity; Immunotherapy; Mice; Needles; Neoplasms; Ovalbumin; T-Lymphocytes, Cytotoxic; Transdermal Patch; Treatment Outcome
PubMed: 31386690
DOI: 10.1371/journal.pone.0220382 -
Journal of Ocular Pharmacology and... Feb 2010An effective treatment modality for posterior eye diseases would provide prolonged delivery of therapeutic agents, including macromolecules, to eye tissues using a safe... (Comparative Study)
Comparative Study
PURPOSE
An effective treatment modality for posterior eye diseases would provide prolonged delivery of therapeutic agents, including macromolecules, to eye tissues using a safe and minimally invasive method. The goal of this study was to assess the ability of a thermosetting gel to deliver a fluorescently labeled protein, Alexa 647 ovalbumin, to the choroid and retina of rats following a single subconjunctival injection of the gel. Additional experiments were performed to compare in vitro to in vivo ovalbumin release rates from the gel.
METHODS
The ovalbumin content of the eye tissues was monitored by spectrophotometric assays of tissue extracts of Alexa 647 ovalbumin from dissected sclera, choroid, and retina at time points ranging from 2 h to 14 days. At the same time points, fluorescence microscopy images of tissue samples were also obtained. Measurement of intact ovalbumin was verified by LDS-PAGE analysis of the tissue extract solutions. In vitro release of Alexa 488 ovalbumin into 37 degrees C PBS solutions from ovalbumin-loaded gel pellets was also monitored over time by spectrophotometric assay. In vivo ovalbumin release rates were determined by measurement of residual ovalbumin extracted from gel pellets removed from rat eyes at various time intervals.
RESULTS
Our results indicate that ovalbumin concentrations can be maintained at measurable levels in the sclera, choroid, and retina of rats for up to 14 days using the thermosetting gel delivery system. The concentration of ovalbumin exhibited a gradient that decreased from sclera to choroid and to retina. The in vitro release rate profiles were similar to the in vivo release profiles.
CONCLUSIONS
Our findings suggest that the thermosetting gel system may be a feasible method for safe and convenient sustained delivery of proteins to choroidal and retinal tissue in the posterior segments of the eye.
Topics: Animals; Choroid; Delayed-Action Preparations; Drug Delivery Systems; Feasibility Studies; Gels; Male; Osmolar Concentration; Ovalbumin; Rats; Rats, Inbred BN; Retina; Sclera; Temperature
PubMed: 20148655
DOI: 10.1089/jop.2009.0059 -
Zeitschrift Fur Naturforschung. C,... 1997We found, by circular dichroism and Raman spectroscopy measurements, that the secondary structure of the native ovalbumin and of its heat-stable form, called... (Comparative Study)
Comparative Study
We found, by circular dichroism and Raman spectroscopy measurements, that the secondary structure of the native ovalbumin and of its heat-stable form, called S-ovalbumin, is a probe of the structural differences between the two proteins. Small angle X-ray scattering and circular dichroism measurements performed on the two proteins under denaturing conditions, with different concentrations of guanidine hydrochloride, show the changes of the tertiary and secondary structure and a different pathway in the unfolding process. These experimental data confirm that the conversion of native ovalbumin into S-ovalbumin is irreversible and reveal that the response of the two proteins to the same chemical environment is different.
Topics: Animals; Chickens; Guanidine; Hot Temperature; Ovalbumin; Protein Conformation; Protein Denaturation; Scattering, Radiation; Spectrum Analysis, Raman; Thermodynamics
PubMed: 9373995
DOI: 10.1515/znc-1997-9-1012 -
Analytical Chemistry Sep 2017Given the wide adoption of polydimethylsiloxane (PDMS) for the rapid fabrication of microfluidic networks and the utility of polyacrylamide gel electrophoresis (PAGE),...
Given the wide adoption of polydimethylsiloxane (PDMS) for the rapid fabrication of microfluidic networks and the utility of polyacrylamide gel electrophoresis (PAGE), we develop a technique for fabrication of PAGE molecular sieving gels in PDMS microchannel networks. In developing the fabrication protocol, we trade-off constraints on materials properties of these two polymer materials: PDMS is permeable to O and the presence of O inhibits the polymerization of polyacrylamide. We present a fabrication method compatible with performing PAGE protein separations in a composite PDMS-glass microdevice, that toggles from an "enclosed" microchannel for PAGE and blotting to an "open" PA gel lane for immunoprobing and readout. To overcome the inhibitory effects of O, we coat the PDMS channel with a 10% benzophenone solution, which quenches the inhibiting effect of O when exposed to UV, resulting in a PAGE-in-PDMS device. We then characterize the PAGE separation performance. Using a ladder of small-to-mid mass proteins (Trypsin Inhibitor (TI); Ovalbumin (OVA); Bovine Serum Albumin (BSA)), we observe resolution of the markers in <60 s, with separation resolution exceeding 1.0 and CVs of 8.4% for BSA-OVA and 2.4% for OVA-TI, with comparable reproducibility to glass microdevice PAGE. We show that benzophenone groups incorporated into the gel through methacrylamide can be UV-activated multiple times to photocapture protein. PDMS microchannel network is reversibly bonded to a glass slide allowing direct access to separated proteins and subsequent in situ diffusion-driven immunoprobing and total protein Sypro red staining. We see this PAGE-in-PDMS fabrication technique as expanding the application and use of microfluidic PAGE without the need for a glass microfabrication infrastructure.
Topics: Adoption; Animals; Cattle; Dimethylpolysiloxanes; Electrophoresis, Polyacrylamide Gel; Immunoblotting; Microfluidic Analytical Techniques; Ovalbumin; Particle Size; Serum Albumin, Bovine; Trypsin Inhibitors
PubMed: 28825964
DOI: 10.1021/acs.analchem.7b02406