Hassan M.E. Azzazy

Gold nanoparticles for molecular diagnostics

Gold nanoparticles (AuNPs) exhibit a unique phenomenon, known as surface plasmon resonance, which is responsible for their large absorption and scattering cross-sections, which are four to five orders of magnitude larger than those of conventional dyes. In addition, their optical properties can be controlled by varying their sizes, shapes and compositions. AuNPs can be easily synthesized and functionalized with different biomolecules including oligonucleotides. Numerous methods have been utilized for detecting AuNPs such as colorimetric, scanometric, fluorescence, surface-enhanced Raman scattering and electrochemical techniques. These unique aspects have permitted the development of novel AuNP-based assays for molecular diagnostics which promise increased sensitivity and specificity, multiplexing capability, and short turnaround times. AuNP-based colorimetric assays in particular show great potential in point-of-care testing assays. This review discusses properties of AuNPs and their utilization for the development of novel molecular assays.

In vitro diagnostic prospects of nanoparticles.

There is a constant need to improve the performance of current diagnostic assays as well as develop innovative testing strategies to meet new testing challenges. The use of nanoparticles promises to help promote in vitro diagnostics to the next level of performance. Quantum dots (QDs), gold nanoparticles (AuNPs), and superparamagnetic nanoparticles are the most promising nanostructures for in vitro diagnostic applications. These nanoparticles can be conjugated to recognition moieties such as antibodies or oligonucleotides for detection of target biomolecules. Nanoparticles have been utilized in immunoassays, immunohistochemistry, DNA diagnostics, bioseparation of specific cell populations, and cellular imaging. Nanoparticle-based diagnostics may open new frontiers for detection of tumours, infectious diseases, bio-terrorism agents, and neurological diseases, to name a few. More work is necessary to fully optimize use of nanoparticles for clinical diagnosis and to resolve some concerns regarding potential health and environmental risks related to their use. However, we envision further developments of nanoparticle-based diagnostics will yield unique assays with enhanced sensitivity and multiplexing capability for the modern clinical laboratory.

Hepatitis B virus genotyping: current methods and clinical implications

Hepatitis B virus (HBV) is a major cause of liver cirrhosis and hepatocellular carcinoma (HCC). Eight different HBV genotypes have been identified with distinct geographical distributions. Different genotyping methods exist including sequencing, INNO-LiPA, restriction fragment polymorphism (RFLP), multiplex PCR, serotyping, oligonucleotide microarray chips, reverse dot blot, restriction fragment mass polymorphism (RFMP), invader assay, and real-time PCR. Several investigators have studied the influence of HBV genotypes on clinical outcomes in chronic HBV patients. This review describes the current genotyping techniques, as well as their advantages and limitations. It also presents the clinical evidence that correlates HBV genotypes to clinical outcomes including disease severity, HCC development, response to therapy, disease chronicity, transplantation outcomes, and occult infection.

Honey/Chitosan Nanofiber Wound Dressing Enriched with Allium sativum and Cleome droserifolia: Enhanced Antimicrobial and Wound Healing Activity

Two natural extracts were loaded within fabricated honey, poly(vinyl alcohol), chitosan nanofibers (HPCS) to develop biocompatible antimicrobial nanofibrous wound dressing. The dried aqueous extract of Cleome droserifolia (CE) and Allium sativum aqueous extract (AE) and their combination were loaded within the HPCS nanofibers in the HPCS-CE, HPCS-AE, and HPCS-AE/CE nanofiber mats, respectively. It was observed that the addition of AE resulted in the least fiber diameter (145 nm), whereas the addition of the AE and CE combination resulted in the least swelling ability and the highest weight loss. In vitro antibacterial testing against Staphylococcus aureus, Escherichia coli, Methicillin-resistant S. aureus (MRSA), and multidrug-resistant Pseudomonas aeruginosa was performed in comparison with the commercial dressing AquacelAg and revealed that the HPCS-AE and HPCS-AE/CE nanofiber mats allowed complete inhibition of S. aureus and the HPCS-AE/CE exhibited mild antibacterial activity against MRSA. A preliminary in vivo study revealed that the developed nanofiber mats enhanced the wound healing process as compared to the untreated control as proved by the enhanced wound closure rates in mice and by the histological examination of the wounds. Moreover, comparison with the commercial dressing Aquacel Ag, the HPCS, and HPCS-AE/CE demonstrated similar effects on the wound healing process, whereas the HPCS/AE allowed an enhanced wound closure rate. Cell culture studies proved the biocompatibility of the developed nanofiber mats in comparison with the commercial Aquacel Ag, which exhibited noticeable cytotoxicity. The developed natural nanofiber mats hold potential as promising biocompatible antibacterial wound dressing.

Optical metal-organic framework sensor for selective discrimination of some toxic metal ions in water

This paper reports the development of a facile and effective approach, based on the use of Zr-based metal-organic frameworks (UiO-66) sensor with micropores geometry, shape and particle morphology for the visual detection and removal of ultra-traces of some toxic metal ions such as Bi(III), Zn(II), Pb(II), Hg(II) and Cd(II). UiO-66 was used as selective carriers for accommodating hydrophobic chromophore probes such as dithizone (DZ) without coupling agent for sensitive and selective discrimination of trace level of toxic analytes. The developed UiO-66 sensor was utilized for the detection of ultra-traces of some toxic metal ions with the naked eye. The new sensor displays high sensitivity and selectivity of a wide range of detectable metals analytes up to 10(-10) mol dm(-3) in solution, in a rapid analyte uptake response (seconds). The developed sensor is stable, cost effective, easy to prepare, and would be useful for rapid detection and removal of ultra-traces of toxic metal ions in water samples.

Cardiac point of care testing: a focused review of current National Academy of Clinical Biochemistry guidelines and measurement platforms.

BACKGROUND Cardiac markers are a cornerstone for assessment of suspected acute coronary syndrome (ACS) patients. The National Academy for Clinical Biochemistry has recently developed practice guidelines for clinical, analytical and point-of-care (POC) testing in the context of ACS. Several technologies have become available for POC applications. SETTING Cardiac troponin is the preferred biochemical marker for diagnosis, risk stratification and guiding management of suspected ACS patients. Samples must be collected with proper timing, typically on presentation and then 6 to 9 h later. Assays should work toward a goal of total CV <10% at the 99th percentile cutpoint, and should adhere to specifications defined by professional organizations. A 1-h or less turnaround time is specified; quantitative POC testing should be implemented if this timing cannot be met consistently by the central laboratory. The same quality criteria for assays apply regardless of testing venue. IMPLEMENTATION/MANAGEMENT: Laboratory medicine personnel must have active role in implementation, choice of technology and management of POC cardiac marker testing. CONCLUSIONS Cardiac troponin measurements at POC are a viable alternative when testing needs cannot be met by the central laboratory. Laboratory medicine must be involved in implementation and ongoing service. Quality of testing must not be compromised by performance at POC.

Unmodified gold nanoparticles for direct and rapid detection of Mycobacterium tuberculosis complex.

OBJECTIVES This work aims to develop rapid nano-gold assay prototypes for specific detection of Mycobacterium tuberculosis complex (MTBC). DESIGN AND METHODS Spherical gold nanoparticles (AuNPs, 14nm) were synthesized by citrate reduction method and characterized by spectrophotometry and SEM. MTB 16s rDNA regions were amplified by PCR and amplicons were detected using genus- and species-specific oligotargeters and AuNPs. In a second prototype, MTBC unamplified genomic DNA was directly detected using species-specific oligo-targeters and AuNPs. RESULTS Detection limits were 1ng for PCR product and 40ng for genomic DNA. The nano-gold prototype detected 45 positive genomic DNA samples which were also positive with automated liquid culture system (BACTEC™ MGIT™) and semi-nested PCR (100% concordance). Following DNA extraction, using standard procedures, the TB nano-gold prototype turnaround time is about 1h. CONCLUSIONS We have developed nano-gold assay prototype for direct and inexpensive detection of MTBC. The developed prototypes are simple, sensitive, rapid and can substitute PCR-based detection. The developed assay may show potential in the clinical diagnosis of TB especially in developing countries.

Gold nanoparticles in the clinical laboratory: principles of preparation and applications

Abstract In order to meet the challenges of effective healthcare, the clinical laboratory is constantly striving to improve testing sensitivity while reducing the required time and cost. Gold nanoparticles (AuNPs) are proposed as one of the most promising tools to meet such goals. They have unique optophysical properties which enable sensitive detection of biomarkers, and are easily amenable to modification for use in different assay formats including immunoassays and molecular assays. Additionally, their preparation is relatively simple and their detection methods are quite versatile. AuNPs are showing substantial promise for effective practical applications and commercial utilization is already underway. This article covers the principles of preparation of AuNPs and their use for development of different diagnostic platforms.

High concentration honey chitosan electrospun nanofibers: Biocompatibility and antibacterial effects

Honey nanofibers represent an attractive formulation with unique medicinal and wound healing advantages. Nanofibers with honey concentrations of <10% were prepared, however, there is a need to prepare nanofibers with higher honey concentrations to increase the antibacterial and wound healing effects. In this work, chitosan and honey (H) were cospun with polyvinyl alcohol (P) allowing the fabrication of nanofibers with high honey concentrations up to 40% and high chitosan concentrations up to 5.5% of the total weight of the fibers using biocompatible solvents (1% acetic acid). The fabricated nanofibers were further chemically crosslinked, by exposure to glutaraldehyde vapor, and physically crosslinked by heating and freezing/thawing. The new HP-chitosan nanofibers showed pronounced antibacterial activity against Staphylococcus aureus but weak antibacterial activity against Escherichia coli. The developed HP-chitosan nanofibers revealed no cytotoxicity effects on cultured fibroblasts. In conclusion, biocompatible, antimicrobial crosslinked honey/polyvinyl alcohol/chitosan nanofibers were developed which hold potential as effective wound dressing.

Challenges in the determination of aminoglycoside antibiotics, a review.

Residues of antibiotics (ABs) in the aquatic environment and in food of animal origin represent a major concern, as prolonged exposure to ABs is a serious health hazard, related to both the side effects of prolonged use and the risk of developing bacterial resistance to various ABs. Given the low levels of the AB residues in complex matrices, the development of sensitive analytical methods represents a major challenge. This is certainly true for the aminoglycoside ABs (AGs) which lack a chromophore and show poor chromatographic properties in reversed-phase liquid chromatography. This paper reviews the current state of the art in the determination of AGs. Attention is paid to extraction, sample clean-up, chromatographic separation, and detection of AGs in both environmental and food samples and in plasma and serum. A general workflow for the analysis of AGs is presented which takes into account the matrix and required level of information.