Tag Archives: PeerRev

Reducing interference with cellular autofluorescence

14 March 2018:

CNBP Research Fellow Nicole Cordina is first author on a new study that reports on two novel methods for reducing interference with cellular autofluorescence for bio-imaging.

Journal: Scientific Reports.

Publication title: Reduced background autofluorescence for cell imaging using nanodiamonds and lanthanide chelates.

Authors: Nicole M. Cordina, Nima Sayyadi, Lindsay M. Parker, Arun Everest-Dass, Louise J. Brown & Nicolle H. Packer.

Abstract:
Bio-imaging is a key technique in tracking and monitoring important biological processes and fundamental biomolecular interactions, however the interference of background autofluorescence with targeted fluorophores is problematic for many bio-imaging applications. This study reports on two novel methods for reducing interference with cellular autofluorescence for bio-imaging. The first method uses fluorescent nanodiamonds (FNDs), containing nitrogen vacancy centers. FNDs emit at near-infrared wavelengths typically higher than most cellular autofluorescence; and when appropriately functionalized, can be used for background-free imaging of targeted biomolecules. The second method uses europium-chelating tags with long fluorescence lifetimes. These europium-chelating tags enhance background-free imaging due to the short fluorescent lifetimes of cellular autofluorescence. In this study, we used both methods to target E-selectin, a transmembrane glycoprotein that is activated by inflammation, to demonstrate background-free fluorescent staining in fixed endothelial cells. Our findings indicate that both FND and Europium based staining can improve fluorescent bio-imaging capabilities by reducing competition with cellular autofluorescence. 30 nm nanodiamonds coated with the E-selectin antibody was found to enable the most sensitive detective of E-selectin in inflamed cells, with a 40-fold increase in intensity detected.

Aptasensors and cytokine detection

10 March 2018:

A new review paper summarising recent advances in aptamer-based biosensors with a specific focus on cytokine sensing has been published in the journal ‘Trends in Analytical Chemistry’. The paper includes CNBP coauthors Fuyuan Zhang, Ewa M.Goldys and Guozhen Liu (pictured).

Journal: Trends in Analytical Chemistry.

Publication title: Advances in Structure-Switching Aptasensing Towards Real Time Detection of Cytokines.

Authors: C. Cao, F. Zhang, E.M. Goldys, G. Liu.

Abstract: Structure-switching aptamer-based biosensors (aptasensors) provide a promising strategy for real-time or near real-time monitoring of analytes in vivo, owing to their reversibility, the versatility of methods available to engineer the aptamer switches, and the ability to tune their dynamic range. Monitoring cell-to-cell communication through cytokine secretions has enormous value in biology and medicine. However, cytokine detection is challenging due to the extremely dynamic, transient cytokine secretion process, and typically low abundances in physiological conditions. Here, we summarise recent advances in structure-switching signaling aptamer-based biosensing with specific focus on cytokine sensing. This Review begins with the survey of cytokine-specific aptamers followed by the designs of elegant sensing platforms based on structure-switching aptamers with different signal readouts such as optic, electrochemistry, and other types. We describe the strategies of signal amplification in aptasensors, and highlight future perspectives of aptasensors for real-time or near real-time detection of cytokines.

Extending depth of field of MOF imaging probes

2 March 2018:

A fully computational method for extending the depth of field of multicore optical fibers (MOF) imagers has been demonstrated by CNBP researchers in a new paper published in the journal ‘Optics Express’. The work shows that the depth of field can be more than doubled for certain spatial frequencies. Lead author on the publication is CNBP Research Fellow Dr Antony Orth from RMIT University.

Journal: Optics Express.

Publication title: Extended depth of field imaging through multicore optical fibers.

Authors: Antony Orth, Martin Ploschner, Ivan S. Maksymov, and Brant C. Gibson.

Abstract: Compact microendoscopes use multicore optical fibers (MOFs) to visualize hard-to-reach regions of the body. These devices typically have a large numerical aperture (NA) and are fixed-focus, leading to blurry images from a shallow depth of field with little focus control. In this work, we demonstrate a method to digitally adjust the collection aperture and therefore extend the depth of field of lensless MOF imaging probes. We show that the depth of field can be more than doubled for certain spatial frequencies, and observe a resolution enhancement of up to 78% at a distance of 50μm from the MOF facet. Our technique enables imaging of complex 3D objects at a comparable working distance to lensed MOFs, but without the requirement of lenses, scan units or transmission matrix calibration. Our approach is implemented in post processing and may be used to improve contrast in any microendoscopic probe utilizing a MOF and incoherent light.

Add-on clip turns smartphone into fully operational microscope

19 February 2018:

Australian researchers from the ARC Centre of Excellence for Nanoscale BioPhotonics (CNBP) have developed a 3D printable ‘clip-on’ that can turn any smartphone into a fully functional microscope.

Reported in the research journal ‘Scientific Reports’, the smartphone microscope is powerful enough to visualise specimens as small as 1/200th of a millimetre, including microscopic organisms, animal and plant cells, blood cells, cell nuclei and more.

The clip-on technology is unique in that it requires no external power or light source to work yet offers high-powered microscopic performance in a robust and mobile handheld package.

And the researchers are making the technology freely available, sharing the 3D printing files publicly so anyone – from scientists to the scientifically curious – can turn their own smartphones into microscopes.

Lead developer and CNBP Research Fellow at RMIT University, Dr Antony Orth (pictured), believes the technology has immense potential as a scientific tool, one that is ideal for use in remote areas and for field-work where larger standalone microscopes are unavailable or impractical.

“We’ve designed a simple mobile phone microscope that takes advantage of the integrated illumination available with nearly all smartphone cameras,” says Dr Orth.

The clip-on has been engineered with internal illumination tunnels that guide light from the camera flash to illuminate the sample from behind. This overcomes issues seen with other microscopy-enabled mobile phone devices says Dr Orth.

“Almost all other phone-based microscopes use externally powered light sources while there’s a perfectly good flash on the phone itself,” he explains. “External LEDs and power sources can make these other systems surprisingly complex, bulky and difficult to assemble.”

“The beauty of our design is that the microscope is useable after one simple assembly step and requires no additional illumination optics, reducing significantly the cost and complexity of assembly. The clip-on is also able to be 3D printed making the device accessible to anyone with basic 3D printing capabilities.”

A further advantage noted by Dr Orth is that the clip-on enables both bright-field and dark-field microscopy techniques to be undertaken. Bright-field microscopy is where a specimen is observed on a bright background. Conversely, dark-field shows the specimen illuminated on a dark background.

“The added dark-field functionality lets us observe samples that are nearly invisible under conventional bright-field operation such as cells in media,” he says. “Having both capabilities in such a small device is extremely beneficial and increases the range of activity that the microscope can be successfully used for.”

Dr Orth believes the potential applications for the smartphone microscope are enormous.

“Our mobile microscope can be used as an inexpensive and portable tool for all types of on-site or remote area monitoring.”

“Water quality, blood samples, environmental observation, early disease detection and diagnosis—these are all areas where our technology can be easily used to good effect.”

Dr Orth sees significant benefit in developing countries for the device.

“Powerful microscopes can be few and far between in some regions,” says Dr Orth. “They’re often only found in larger population centres and not in remote or smaller communities. Yet their use in these areas can be essential—for determining water quality for drinking, through to analysing blood samples for parasites, or for disease diagnosis including malaria.”

To ensure that this technology can be utilised the world over, the files for the 3D printing of the microscope clip-on are being made freely available. They are available for download at the CNBP web site – http://cnbp.org.au/online-tools.

“Ideally, a phone microscope should take advantage of the integrated flash found in nearly every modern mobile, avoiding the need for external lighting and power. It should also be as compact and easy to assemble as possible. It is this design philosophy that inspired us in the development of this add-on clip,” says Dr Orth.

The new phone microscope has already been tested by Dr Orth and his CNBP colleagues in a number of areas, successfully visualizing samples ranging from cell culture, to zooplankton to live cattle semen in support of livestock fertility testing.

Below: Cells being viewed by an add-on clip that turns a smartphone into a fully operational microscope.

Detonation nanodiamonds to aid bioimaging

6 February 2018:

Tiny 5 nm detonation nanodiamonds glow in different colors and their fluorescence is pH dependent, reports a new paper by CNBP scientists published today in the Nature journal Scientific Reports.

Lead author of the paper Dr Philipp Reineck from RMIT University (Former CNBP Research Fellow and current CNBP Associate Investigator) notes that the research is particulalry exciting as the fluorescence lifetime of the detonation nanodiamonds makes fluorescence lifetime imaging (FLIM) for bioimaging applications feasible.

Journal: Scientific Reports.

Publication title: Visible to near-IR fluorescence from single-digit detonation nanodiamonds: excitation wavelength and pH dependence.

Authors: Philipp Reineck, Desmond W. M. Lau, Emma R. Wilson, Nicholas Nunn, Olga A. Shenderova & Brant C. Gibson.

Abstract: Detonation nanodiamonds are of vital significance to many areas of science and technology. However, their fluorescence properties have rarely been explored for applications and remain poorly understood. We demonstrate significant fluorescence from the visible to near-infrared spectral regions from deaggregated, single-digit detonation nanodiamonds dispersed in water produced via post-synthesis oxidation. The excitation wavelength dependence of this fluorescence is analyzed in the spectral region from 400 nm to 700 nm as well as the particles’ absorption characteristics. We report a strong pH dependence of the fluorescence and compare our results to the pH dependent fluorescence of aromatic hydrocarbons. Our results significantly contribute to the current understanding of the fluorescence of carbon-based nanomaterials in general and detonation nanodiamonds in particular.

Diabetes and early pregnancy

1 February 2018:

CNBP and Robinson Research Institute researcher Dr Hannah Brown, University of Adelaide is lead author on a newly published paper that looks to understand why pregnancy failure and pregnancy loss occurs in women with diabetes. The paper was published in the Nature journal Scientific Reports.

Publication titlePericonception onset diabetes is associated with embryopathy and fetal growth retardation, reproductive tract hyperglycosylation and impaired immune adaptation to pregnancy.

Authors: Hannah M. Brown, Ella S. Green, Tiffany C. Y. Tan, Macarena B. Gonzalez, Alice R. Rumbold, M. Louise Hull, Robert J. Norman, Nicolle H. Packer, Sarah A. Robertson & Jeremy G. Thompson.

Abstract: Diabetes has been linked with impaired fertility but the underlying mechanisms are not well defined. Here we use a streptozotocin-induced diabetes mouse model to investigate the cellular and biochemical changes in conceptus and maternal tissues that accompany hyperglycaemia. We report that streptozotocin treatment before conception induces profound intra-cellular protein β-O-glycosylation (O-GlcNAc) in the oviduct and uterine epithelium, prominent in early pregnancy. Diabetic mice have impaired blastocyst development and reduced embryo implantation rates, and delayed mid-gestation growth and development. Peri-conception changes are accompanied by increased expression of pro-inflammatory cytokine Trail, and a trend towards increased Il1a, Tnf and Ifng in the uterus, and changes in local T-cell dynamics that skew the adaptive immune response to pregnancy, resulting in 60% fewer anti-inflammatory regulatory T-cells within the uterus-draining lymph nodes. Activation of the heat shock chaperones, a mechanism for stress deflection, was evident in the reproductive tract. Additionally, we show that the embryo exhibits elevated hyper-O-GlcNAcylation of both cytoplasmic and nuclear proteins, associated with activation of DNA damage (ɣH2AX) pathways. These results advance understanding of the impact of peri-conception diabetes, and provide a foundation for designing interventions to support healthy conception without propagation of disease legacy to offspring.

Biological hydrogen peroxide detection

31 January 2018:

Two important sensing architectures for detecting hydrogen peroxide, aryl boronates and benzils, have been compared by CNBP researchers, using novel boron-dipyrromethene (BODIPY) fluorescent probes. Lead author of the publication was Dr Malcolm Purdey (pictured).

Publication titleBiological hydrogen peroxide detection with aryl boronate and benzil BODIPY-based fluorescent probes.

Journal: Sensors and Actuators B: Chemical.

Authors: Malcolm S. Purdey, Hanna J. McLennan, Melanie L. Sutton-McDowall, Daniel W. Drumm, Xiaozhou Zhang, Patrick K. Capon, Sabrina Heng, Jeremy G. Thompson, Andrew D. Abell.

Abstract: The detection of hydrogen peroxide (H2O2) using fluorescent probes is critical to the study of oxidative stress in biological environments. Two important sensing architectures for detecting H2O2, aryl boronates and benzils, are compared here using novel boron-dipyrromethene (BODIPY) fluorescent probes. The aryl boronate PeroxyBODIPY-1 (PB1) and benzil-based nitrobenzoylBODIPY (NbzB) were synthesised from a common BODIPY intermediate in order to compare sensitivity and selectivity to H2O2. The aryl boronate PB1 gives the highest change in fluorescence on reaction with H2O2 while the benzil NbzB exhibits exclusive selectivity for H2O2 over other reactive oxygen species (ROS). Both proved to be cell-permeable, with PB1 being able to detect H2O2 in denuded bovine oocytes. The strengths of these aryl boronate and benzil probes can now be exploited concurrently to elucidate biological mechanisms of H2O2 production and oxidative stress.

Magnetically sensitive optical fibre demonstrated

19 January 2018:

A new paper featuring CNBP researchers demonstrates magnetically sensitive nanodiamond-doped tellurite glass fibres. This work is a first step towards magneto-sensitive fibre devices which could be used in medical magneto-endoscopy and remote mineral exploration sensing. First author of the paper is CNBP AI, Dr Yinlan Ruan from the University of Adelaide.

Journal: Scientific Reports.

Publication titleMagnetically sensitive nanodiamond-doped tellurite glass fibers.

Authors: Yinlan Ruan, David A. Simpson, Jan Jeske, Heike Ebendorff-Heidepriem, Desmond W. M. Lau, Hong Ji, Brett C. Johnson, Takeshi Ohshima, Shahraam Afshar V., Lloyd Hollenberg, Andrew D. Greentree, Tanya M. Monro & Brant C. Gibson.

Abstract: Traditional optical fibers are insensitive to magnetic fields, however many applications would benefit from fiber-based magnetometry devices. In this work, we demonstrate a magnetically sensitive optical fiber by doping nanodiamonds containing nitrogen vacancy centers into tellurite glass fibers. The fabrication process provides a robust and isolated sensing platform as the magnetic sensors are fixed in the tellurite glass matrix. Using optically detected magnetic resonance from the doped nanodiamonds, we demonstrate detection of local magnetic fields via side excitation and longitudinal collection. This is a first step towards intrinsically magneto-sensitive fiber devices with future applications in medical magneto-endoscopy and remote mineral exploration sensing.

New aptasensor able to detect intracellular IFN-γ

8 January 2018:

A new research paper, resulting from a partnership between CNBP and Jilin University has been published, reporting on the development of a new aptasensor which is able to detect ultrasmall concentrations of intracellular IFN-γ. This simple and highly sensitive sensor is able to be used for real-time bio-imaging, providing  a universal sensing platform for monitoring a spectrum of molecules secreted by cells.

Journal: ACS Sensors.

Publication title: “Turn-on” Fluorescent Aptasensor Based on AIEgen Labeling for the Localization of IFN-γ in Live Cells.

Author: Ke Ma, Fengli Zhang, Nima Sayyadi, Wenjie Chen, Ayad G. Anwer, Andrew Care, Bin Xu , Wenjing Tian, Ewa M. Goldys and Guozhen Liu.

Abstract: We report an aggregation-induced emission fluorogen (AIEgen)-based turn-on fluorescent aptasensor able to detect the ultrasmall concentration of intracellular IFN-γ. The aptasensor consists of an IFN-γ aptamer labeled with a fluorogen with a typical aggregation-induced emission (AIE) characteristic, which shows strong red emission only in the presence of IFN-γ. The aptasensor is able to effectively monitor intracellular IFN-γ secretion with the lowest detection limit of 2 pg mL-1, and it is capable of localizing IFN-γ in live cells during secretion, with excellent cellular permeability and biocompatibility as well as low cytotoxicity. This probe is able to localize the intracellular IFN-γ at a low concentration <10 pg mL-1, and it is successfully used for real-time bioimaging. This simple and highly sensitive sensor may enable the exploration of cytokine pathways and their dynamic secretion process in the cellular environment. It provides a universal sensing platform for monitoring a spectrum of molecules secreted by cells.

Multi-LED light source for hyperspectral imaging

15 December 2017:

A multi-LED light source has been investigated by CNBP researchers for use in hyperspectral imaging. The research featured in the journal ‘Optics Express’  with the lead author on the paper, PhD student Kashif Islam, based at Macquarie University.

Journal: Optics Express.

Publication titleMulti-LED light source for hyperspectral imaging.

Author: Kashif Islam, Martin Ploschner, and Ewa M. Goldys.

Abstract: We investigate a novel light source design for efficient collection of UV light from multiple LEDs. The design is based on a truncated cone with a large circular lid incorporating LEDs on one side, and a small circular exit aperture (diameter of 9 mm) on the other side. The exit aperture size allows a simple coupling with secondary optics of a microscope for hyperspectral imaging that requires a wide spectrum of frequencies of illuminating light. The light collection efficiency was optimized with respect to cone length and diameter of the LED lid. In all simulations, we use a highly UV-reflecting aluminum coating on the inner surfaces of the cone. The influence of the LED positions on the cone efficiency is determined by varying the LED distance from the central axis as well as the interLED distance. We found the maximum efficiency of the cone is realized for LEDs positioned at the center, and the shorter is the inter-LED distance, the better is the performance of the light source.