Gold Nanoparticles For Biomolecular Assays

The amalgamation of nanotechnology and biology has led to the development ofnew types of hybrid materials that are expected to produce major advances in areas such as materials science, therapeutics and diagnostics. One of the most promising developments is the use ofnanoparticles (NPs) as labels for the detection of analytes in biological assays. The aim of this research project was to prepare gold nanoparticle (GNP) labels for use in such assays. In chapter 1, the optical properties and the use of GNPs in homogeneous and heterogeneous colorimetric assays are reviewed. In chapter 2 a simple conjugation method is introduced that not only allows almost any biological molecule or hapten to be attached to GNPs but also allows the user to control or vary the mean number of molecules per particle. In this method a high molecular weight aminodextran polymer is functionalized with the molecule of choice and chemical attachment groups that are used to covalently anchor the polymer to the GNPs. This method was used to conjugate biotin and 1125 functionalized dextrans to GNPs. These functionalized dextrans were then used to investigate the conjugation procedure in more_detail. Results from GNP titrations and microbead assays demonstrate that the minimum amount of functionalized dextran required to prevent salt-induced flocculation ofthe GNPs (equivalence point) is the amount required to coat all of the GNPs and at this point there is no free functionalized dextran in solution. In chapter 3 the described method was used to conjugate different numbers DNP haptens to GNPs and then these labels were used in non-traditional reagent-limited lateral flow immunoassays. The number of molecules per GNP is varied by simply adjusting the stoichiometry of reagents in the dextran functionalization reaction. Controlling the number of molecules per particle can have important consequences on the sensitivity of a biological assay. Results showed that when the number of DNP molecules per particle decreased, there was an increase in the sensitivity of the assay. Furthermore when the results from these immunoassays were compared to those obtained from traditional reagent-limited lateral flow immunoassays, the nontraditional format proved to be over 50 % more sensitive. In chapter 4 the conjugation method was used to attach oligonucleotides to GNPs for use in a nucleic acids lateral flow (NALF) device. Although NALF devices are available commercially, detection is usually achieved with the use of antibodies or haptens which can be both problematic and expensive. In addition, many of these devices have issues with sensitivity and are often interfaced with complicated target amplification / purification protocols. In chapter 4 an antibody / hapten independent NALF device is described that can be used to detect the un-purified products from a simple polymerase chain reaction (PCR) amplification protocol. Using the developed NALF device it was possible to detect specific amplification products corresponding to ~1 attomole oftemplate molecules with the unaided eye

Dairy-inspired coatings for bone implants from whey protein isolate-derived self-assembled fibrils

To improve integration of a biomaterial with surrounding tissue, its surface properties may be modified by adsorption of biomacromolecules, e.g. fibrils. Whey protein isolate (WPI), a dairy industry by-product, supports osteoblastic cell growth. WPI’s main component, β-lactoglobulin, forms fibrils in acidic solutions. In this study, aiming to develop coatings for biomaterials for bone contact, substrates were coated with WPI fibrils obtained at pH 2 or 3.5. Importantly, WPI fibrils coatings withstood autoclave sterilization and appeared to promote human bone marrow stromal cells (hBMSC) spreading and differentiation. In the future, WPI fibrils coatings could facilitate immobilization of biomolecules with growth stimulating or antimicrobial propertie

Effect of Polymer Demixed Nanotopographies on Bacterial Adhesion and Biofilm Formation

As the current global threat of antimicrobial resistance (AMR) persists, developing alternatives to antibiotics that are less susceptible to resistance is becoming an urgent necessity. Recent advances in biomaterials have allowed for the development and fabrication of materials with discrete surface nanotopographies that can deter bacteria from adhering to their surface. Using binary polymer blends of polystyrene (PS), poly(methyl methacrylate) (PMMA) and polycaprolactone (PCL) and varying their relative concentrations, PS/PCL, PS/PMMA and PCL/PMMA polymer demixed thin films were developed with nanoisland, nanoribbon and nanopit topographies. In the PS/PCL system, PS segregates to the air-polymer interface, with the lower solubility PCL preferring the substrate-polymer interface. In the PS/PMMA and PCL/PMMA systems, PMMA prefers the air-polymer interface due to its greater solubility and lower surface energy. The anti-adhesion efficacy of the demixed films were tested against Pseudomonas aeruginosa (PA14). PS/PCL and PCL/PMMA demixed films showed a significant reduction in cell counts adhered on their surfaces compared to pure polymer control films, while no reduction was observed in the counts adhered on PS/PMMA demixed films. While the specific morphology did not affect the adhesion, a relationship between bacterial cell and topographical surface feature size was apparent. If the surface feature was smaller than the cell, then an anti-adhesion effect was observed; if the surface feature was larger than the cell, then the bacteria preferred to adhere. View Full-Tex

A randomised controlled trial linking mental health inpatients to community smoking cessation supports: A study protocol

<p>Abstract</p> <p>Background</p> <p>Mental health inpatients smoke at higher rates than the general population and are disproportionately affected by tobacco dependence. Despite the advent of smoke free policies within mental health hospitals, limited systems are in place to support a cessation attempt post hospitalisation, and international evidence suggests that most smokers return to pre-admission smoking levels following discharge. This protocol describes a randomised controlled trial that will test the feasibility, acceptability and efficacy of linking inpatient smoking care with ongoing community cessation support for smokers with a mental illness.</p> <p>Methods/Design</p> <p>This study will be conducted as a randomised controlled trial. 200 smokers with an acute mental illness will be recruited from a large inpatient mental health facility. Participants will complete a baseline survey and will be randomised to either a multimodal smoking cessation intervention or provided with hospital smoking care only. Randomisation will be stratified by diagnosis (psychotic, non-psychotic). Intervention participants will be provided with a brief motivational interview in the inpatient setting and options of ongoing smoking cessation support post discharge: nicotine replacement therapy (NRT); referral to Quitline; smoking cessation groups; and fortnightly telephone support. Outcome data, including cigarettes smoked per day, quit attempts, and self-reported 7-day point prevalence abstinence (validated by exhaled carbon monoxide), will be collected via blind interview at one week, two months, four months and six months post discharge. Process information will also be collected, including the use of cessation supports and cost of the intervention.</p> <p>Discussion</p> <p>This study will provide comprehensive data on the potential of an integrated, multimodal smoking cessation intervention for persons with an acute mental illness, linking inpatient with community cessation support.</p> <p>Trial Registration</p> <p>Australian and New Zealand Clinical Trials Registry ANZTCN: <a href="http://www.anzctr.org.au/ACTRN12609000465257.aspx">ACTRN12609000465257</a></p

Gold Nanoparticles For Biomolecular Assays

The amalgamation of nanotechnology and biology has led to the development ofnew types of hybrid materials that are expected to produce major advances in areas such as materials science, therapeutics and diagnostics. One of the most promising developments is the use ofnanoparticles (NPs) as labels for the detection of analytes in biological assays. The aim of this research project was to prepare gold nanoparticle (GNP) labels for use in such assays. In chapter 1, the optical properties and the use of GNPs in homogeneous and heterogeneous colorimetric assays are reviewed. In chapter 2 a simple conjugation method is introduced that not only allows almost any biological molecule or hapten to be attached to GNPs but also allows the user to control or vary the mean number of molecules per particle. In this method a high molecular weight aminodextran polymer is functionalized with the molecule of choice and chemical attachment groups that are used to covalently anchor the polymer to the GNPs. This method was used to conjugate biotin and 1125 functionalized dextrans to GNPs. These functionalized dextrans were then used to investigate the conjugation procedure in more_detail. Results from GNP titrations and microbead assays demonstrate that the minimum amount of functionalized dextran required to prevent salt-induced flocculation ofthe GNPs (equivalence point) is the amount required to coat all of the GNPs and at this point there is no free functionalized dextran in solution. In chapter 3 the described method was used to conjugate different numbers DNP haptens to GNPs and then these labels were used in non-traditional reagent-limited lateral flow immunoassays. The number of molecules per GNP is varied by simply adjusting the stoichiometry of reagents in the dextran functionalization reaction. Controlling the number of molecules per particle can have important consequences on the sensitivity of a biological assay. Results showed that when the number of DNP molecules per particle decreased, there was an increase in the sensitivity of the assay. Furthermore when the results from these immunoassays were compared to those obtained from traditional reagent-limited lateral flow immunoassays, the nontraditional format proved to be over 50 % more sensitive. In chapter 4 the conjugation method was used to attach oligonucleotides to GNPs for use in a nucleic acids lateral flow (NALF) device. Although NALF devices are available commercially, detection is usually achieved with the use of antibodies or haptens which can be both problematic and expensive. In addition, many of these devices have issues with sensitivity and are often interfaced with complicated target amplification / purification protocols. In chapter 4 an antibody / hapten independent NALF device is described that can be used to detect the un-purified products from a simple polymerase chain reaction (PCR) amplification protocol. Using the developed NALF device it was possible to detect specific amplification products corresponding to ~1 attomole oftemplate molecules with the unaided eye.EThOS - Electronic Theses Online ServiceGBUnited Kingdo

Nitric Oxide Releasing Polymeric Coatings for the Prevention of Biofilm Formation

The ability of nitric oxide (NO)-releasing polymer coatings to prevent biofilm formation is described. NO-releasing coatings on (poly(ethylene terephthalate) (PET) and silicone elastomer (SE)) were fabricated using aminosilane precursors. Pristine PET and SE were oxygen plasma treated, followed by immobilisation of two aminosilane molecules: N-(3-(trimethoxysilyl)propyl)diethylenetriamine (DET3) and N-(3-trimethoxysilyl)propyl)aniline (PTMSPA). N-diazeniumdiolate nitric oxide donors were formed at the secondary amine sites on the aminosilane molecules producing NO-releasing polymeric coatings. The NO payload and release were controlled by the aminosilane precursor, as DET3 has two secondary amine sites and PTMSPA only one. The antibacterial efficacy of these coatings was tested using a clinical isolate of Pseudomonas aeruginosa (PA14). All NO-releasing coatings in this study were shown to significantly reduce P. aeruginosa adhesion over 24 h with the efficacy being a function of the aminosilane modification and the underlying substrate. These NO-releasing polymers demonstrate the potential and utility of this facile coating technique for preventing biofilms for indwelling medical devices

Greater Attendance at a Community Weight Loss Programme over the First 12 Weeks Predicts Weight Loss at 2 Years.

BACKGROUND: There is considerable heterogeneity in long-term weight loss among people referred to obesity treatment programmes. It is unclear whether attendance at face-to-face sessions in the early weeks of the programme is an independent predictor of long-term success. OBJECTIVE: To investigate whether frequency of attendance at a community weight loss programme over the first 12 weeks is associated with long-term weight change. METHODS: Participants were randomised to receive brief support only (control, n = 211), or a weight loss programme for 12 weeks (n = 530) or 52 weeks (n = 528). This study included participants with data on session attendance over the first 12 weeks (n = 889) compared to the control group. The association between attendance (continuously) and weight loss was explored using a linear model. A multi-level mixed-effects linear model was used to investigate whether attendance (categorised as 0, 1, 2-5, 6-9, and 10-12 sessions) was associated with weight loss at 3, 12, and 24 months compared to the control. RESULTS: For every session attended in the first 12 weeks, the average weight loss was -0.259 kg/session at 24 months (p = 0.005). Analysis by attendance group found only those attending 10-12 sessions had significantly greater weight loss (-7.5 kg [95% CI -8.1 to -6.9] at 12 months; -4.7 kg [95% CI -5.3 to -4.1] at 24 months) compared to the control group (-3.4 [95% CI -4.5 to -2.4] at 12 months, -2.5 [95% CI -3.5 to -1.5] at 24 months). Early attendance was higher for people ≥70 years, but there was no evidence of a difference by gender, ethnicity, education, or income. CONCLUSIONS: Greater attendance at a community weight loss programme in the first 12 weeks is associated with enhanced weight loss up to 24 months. Regular attendance at a programme could be used as a criterion for continued provision of weight loss services to maximise the cost-effectiveness of interventions.The WRAP trial is funded by the National Prevention Research Initiative through research grant MR/J000493/1. The funding partners relevant to this award are (in alphabetical order): Alzheimer’s Research Trust, Alzheimer’s Society, Biotechnology and Biological Sciences Research Council, British Heart Foundation, Cancer Research UK, Chief Scientist Office, Scottish Government Health Directorate, Department of Health, Diabetes 15 UK, Economic and Social Research Council, Health and Social Care Research and Development Division of the Public Health Agency (HSC R&D Division), UK Medical Research Council (MRC), The Stroke Association, Welcome Trust, Welsh Assembly Government, and World Cancer Research Fund. The cost of the Weight Watchers programme was funded by WW (formally Weight Watchers International) as part of an MRC Industrial Collaboration Award. During the conduct of the WRAP trial SAJ and ALA were supported by the MRC (grant number U105960389). SAJ and PA are currently NIHR Senior Investigators and supported by the Oxford NIHR Biomedical Research Centre and Oxford NIHR Collaboration and Leadership in Applied Health Research and Care (CLAHRC). CP’s time on this project is also supported by the Oxford NIHR CLAHRC. ALA is supported by the MRC (grant MC_UU_12015/4)