Browsing by Author "Du Preez, Ilse"

Now showing 1 - 6 of 6
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    Application of monoclonal anti-mycolate antibodies in serological diagnosis of tuberculosis
    (2024-11) Truyts, Alma E; Du Preez, Ilse; Maesela, Maushe E; Scriba, Manfred R; Baillie, L; Jones, AT; Land, KJ; Verschoor, JA; Lemmer, Yolandy
    Patient loss to follow-up caused by centralised and expensive diagnostics that are reliant on sputum is a major obstacle in the fight to end tuberculosis. An affordable, non-sputum biomarker-based, point-of-care deployable test is needed to address this. Serum antibodies binding the mycobacterial cell wall lipids, mycolic acids, have shown promise as biomarkers for active tuberculosis. However, anti-lipid antibodies are of low affinity, making them difficult to detect in a lateral flow immunoassay—a technology widely deployed at the point-of-care. Previously, recombinant monoclonal anti-mycolate antibodies were developed and applied to characterise the antigenicity of mycolic acid. We now demonstrate that these anti-mycolate antibodies specifically detect hexane extracts of mycobacteria. Secondary antibody-mediated detection was applied to detect the displacement of the monoclonal mycolate antibodies by the anti-mycolic acid antibodies present in tuberculosis-positive guinea pig and human serum samples. These data establish proof-of-concept for a novel lateral flow immunoassay for tuberculosis provisionally named MALIA—mycolate antibody lateral flow immunoassay.
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    Development of bioconjugated dye-doped poly(styrene-co-maleimide) nanoparticles as a new bioprobe
    (Royal Society of Chemistry, 2015-02) Swanepoel, Andri; Du Preez, Ilse; Mahlangu, T; Chetty, A; Klumperman, B
    Fluorescent dye-doped poly(styrene-co-maleimide) nanoparticles were synthesised and functionalized with amine groups and avidin and the dye-loading was varied. The nanoparticles showed great potential as bioprobes when conjugated with biotinylated antibodies
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    hFOB 1.19 osteoblast cells grown on a biomimetic biphasic nanoscaffold: an in vitro evaluation for possible bone tissue engineering
    (Allied Academies, 2018) Du Preez, Ilse; Richter, Wim; Van Papendorp, D; Joubert, A
    Much research over the past five decades has focussed on the repair and replacement of bone. Recently, the research focus has shifted to nanotechnology since it provides a platform from which to alter and possibly improve materials’ properties. In this study we have made use of previously developed electrospun biphasic nanoscaffolds to culture osteoblast cells on, and investigate specific responses of the cells towards the scaffolds. Osteoclast-like cells and osteoblast cells were cultured separately on the nanoscaffolds and the proliferation, adhesion and cellular response were determined. In this study, the mineralisation of the osteoblast cells was observed in a time study. The intracellular calcium ion concentration and nitric oxide concentration were determined in vitro while the cells were proliferating on the scaffolds. The expression of endothelial and inducible nitric oxide synthase was determined immunohistochemically. Quantitative data were obtained from fluorometer studies. Qualitative data was supplied by light- and fluorescent confocal microscopy. During studies with microscopy, a minimum of five representative images from each sample were captured. The cells showed increased mineralisation over time. An increase in intracellular Ca2+ was not observed when compared to the controls. However, an increase in intracellular nitric oxide formation was detected. Expression of endothelial nitric oxide synthase but not inducible nitric oxide synthase was detected in vitro. From the results we can conclude that the scaffolds are biocompatible and conducive to healthy cell growth and differentiation and could possibly be applied in non-load-bearing bone regeneration and repair applications.
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    Immunogenic profile of a plant-produced nonavalent African horse sickness viral protein 2 (VP2) vaccine in IFNAR-/-mice
    (2024-04) O’Kennedy, Martha M; Roth, Robyn; Ebersohn, K; Du Plessis, LH; Mamputha, Sipho; Rutkowska, Daria A; Du Preez, Ilse; Verschoor, JA; Lemmer, Yolandy
    A safe, highly immunogenic multivalent vaccine to protect against all nine serotypes of African horse sickness virus (AHSV), will revolutionise the AHS vaccine industry in endemic countries and beyond. Plant-produced AHS virus-like particles (VLPs) and soluble viral protein 2 (VP2) vaccine candidates were developed that have the potential to protect against all nine serotypes but can equally well be formulated as mono- and bi-valent formulations for localised outbreaks of specific serotypes. In the first interferon a/ß receptor knock-out (IFNAR-/-) mice trial conducted, a nine-serotype (nonavalent) vaccine administered as two pentavalent (5 µg per serotype) vaccines (VLP/VP2 combination or exclusively VP2), were directly compared to the commercially available AHS live attenuated vaccine. In a follow up trial, mice were vaccinated with an adjuvanted nine-serotype multivalent VP2 vaccine in a prime boost strategy and resulted in the desired neutralising antibody titres of 1:320, previously demonstrated to confer protective immunity in IFNAR-/- mice. In addition, the plant-produced VP2 vaccine performed favourably when compared to the commercial vaccine. Here we provide compelling data for a nonavalent VP2-based vaccine candidate, with the VP2 from each serotype being antigenically distinguishable based on LC-MS/MS and ELISA data. This is the first preclinical trial demonstrating the ability of an adjuvanted nonavalent cocktail of soluble, plant-expressed AHS VP2 proteins administered in a prime-boost strategy eliciting high antibody titres against all 9 AHSV serotypes. Furthermore, elevated T helper cells 2 (Th2) and Th1, indicative of humoral and cell-mediated memory T cell immune responses, respectively, were detected in mouse serum collected 14 days after the multivalent prime-boost vaccination. Both Th2 and Th1 may play a role to confer protective immunity. These preclinical immunogenicity studies paved the way to test the safety and protective efficacy of the plant-produced nonavalent VP2 vaccine candidate in the target animals, horses.
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    Protective efficacy of a plant-produced beta variant rSARS-CoV-2 VLP vaccine in golden Syrian hamsters
    (2024-02) Lemmer, Yolandy; Chapman, R; Abolnik, C; Smith, Tanja; Schäfer, G; Hermanus, T; Du Preez, Ilse; Sepotokele, Kamogelo M; Roth, Robyn; Truyts, Alma; O'Kennedy, Maretha M
    In the quest for heightened protection against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants, we engineered a prototype vaccine utilizing the plant expression system of Nicotiana benthamiana, to produce a recombinant SARS-CoV-2 virus-like particle (VLP) vaccine presenting the S-protein from the Beta (B.1.351) variant of concern (VOC). This innovative vaccine, formulated with either a squalene oil-in-water emulsion or a synthetic CpG oligodeoxynucleotide adjuvant, demonstrated efficacy in a golden Syrian Hamster challenge model. The Beta VLP vaccine induced a robust humoral immune response, with serum exhibiting neutralization not only against SARS-CoV-2 Beta but also cross-neutralizing Delta and Omicron pseudoviruses. Protective efficacy was demonstrated, evidenced by reduced viral RNA copies and mitigated weight loss and lung damage compared to controls. This compelling data instills confidence in the creation of a versatile platform for the local manufacturing of potential pan-sarbecovirus vaccines, against evolving viral threats.
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    Protective immunity of plant-produced African horse sickness virus serotype 5 chimaeric virus-like particles (VLPs) and viral protein 2 (VP2) vaccines in IFNAR-/- mice
    (2022-08) O'Kennedy, Maretha M; Coetzee, P; Koekemoer, O; Du Plessis, L; Lourens, CW; Kwezi, Lusisizwe; Du Preez, Ilse; Mamputha, Sipho; Rutkowska, Daria A; Lemmer, Yolandy
    Next generation vaccines have the capability to contribute to and revolutionise the veterinary vaccine industry. African horse sickness (AHS) is caused by an arbovirus infection and is characterised by respiratory distress and/or cardiovascular failure and is lethal to horses. Mandatory annual vaccination in endemic areas curtails disease occurrence and severity. However, development of a next generation AHSV vaccine, which is both safe and efficacious, has been an objective globally for years. In this study, both AHSV serotype 5 chimaeric virus-like particles (VLPs) and soluble viral protein 2 (VP2) were successfully produced in Nicotiana benthamiana ΔXT/FT plants, partially purified and validated by gel electrophoresis, transmission electron microscopy and liquid chromatography-mass spectrometry (LC-MS/MS) based peptide sequencing before vaccine formulation. IFNAR-/- mice vaccinated with the adjuvanted VLPs or VP2 antigens in a 10 µg prime-boost regime resulted in high titres of antibodies confirmed by both serum neutralising tests (SNTs) and enzyme-linked immunosorbent assays (ELISA). Although previous studies reported high titres of antibodies in horses when vaccinated with plant-produced AHS homogenous VLPs, this is the first study demonstrating the protective efficacy of both AHSV serotype 5 chimaeric VLPs and soluble AHSV-5 VP2 as vaccine candidates. Complementary to this, coating ELISA plates with the soluble VP2 has the potential to underpin serotype-specific serological assays.