Facilities within the Diabetes & Islet Biology Group laboratories at Western Sydney University School of Medicine.
Olink Signature Q100
The Olink Signature Q100 instrument is the latest addition to the Hardikar lab's capabilities, further advancing our quest to decipher multi-omics dynamic biomarker signatures. This system has been procured to add on to existing microRNA, cfDNA, and metabolite (GC/MS) capabilities that our lab group already has. Additional facilities and support in multi-omic technologies is available through the Mass Spectrometry Facility in our building. For more details on the proprietary Olink technology in our lab, including instrument capabilities and proteomics and cytokine panels, please visit https://olink.com/instruments/olink-signature-q100
Smart USSING
SmartUSSING is a better way to increase the impact of your Ussing experiments by adding electrophysiological data. With an easy-to-use design, user-friendly software, and a smaller footprint, SmartUSSING makes electrophysiology easy and accessible for anyone. This instrument has been procured through the Novo Nordisk Foundation grant and is mainly used to measure the transepithelial transport of metabolites and small RNA moleules across human gut cells. For more information on the system, please visit: https://www.warneronline.com/SmartUSSING
QiaCube - HT Robotic RNA/DNA isolation system
All of the RNA and DNA isolation is preferably carried out on the QiaCube-HT high throughput robotic system. This offers the capacity to isolate 96 samples from a single run and yields CVs less than 5% between different plates. The system is optimized to use TriZol-based protocols on the QiaCube system and enables robotic-based isolation of nucleic acids from biofluids. This instrument was procured through funding support from the JDRF Australia - Helmsley Trust research project to AAH.
Nanodrop Lite Spectrophotometer ("Eeny")
The Thermo Scientific™ NanoDrop Lite offers a full-spectrum, UV-Vis spectrum to quantify and assess purity of DNA and RNA (we do not use this spectrophotometer for protein). The NanoDrop Lite is one of the first microvolume spectrophotometers with patented sample retention technology that measures sample volumes as small as 1μL. This system is dedicated for purified protein concentration assessment for a quick screen at 280nm and other Spectroscopy, Elemental & Isotope Analysis.
Nanodrop Eight ("Meeny")
The Thermo Scientific™ NanoDrop™ Eight is a UV-Visible spectrophotometer developed for micro-volume analysis of a wide variety of analytes. The patented sample retention system enables the measurement of highly concentrated samples without the need for dilutions. The instrument accurately measures up to 8 individual 1 µl samples in one measurement cycle. This system is dedicated for quick nucleic acid assessment in the lab. Please note - a Qubit fluorimeter is available across from our lab for specific applications.
QIAxpert ("Miny")
This is an innovative high-speed microfluidic UV/VIS spectrophotometer with capacity to differentiate between DNA, RNA and sample impurities, processing up to 16 samples in less than two minutes using a contamination-free nanofluidic slide (consumables required). Once loaded the sample remains saved up to two hours as there is no evaporation loss. Since conventional UV/Vis measurement relies on the absorbance at 260 nm (both DNA and RNA can absorb at this wavelength resulting in an overestimation of nucleic acid concentration). QIAxpert contains a wide range of pre-installed methods which can measure protein concentration via absorbance or colorimetric assay, monitor bacterial growth, display results on a smart device via QR code, can calibrate by itself and can generate the results in multiple formats.
QIAxcel Connect ("Moe")
Fully automate the fragment analysis and quality control of up to 96 samples per run. This capillary-electrophoresis instrument uses ready-to-run gel cartridges that eliminate tedious gel or consumable preparations reducing handling errors. This system is dedicated to QC check for RNA/DNA samples, library preperation, High-resolution genotyping, Fast single or multiplex PCR screening, QC of cell-free DNA, NGS library QC, Fast single-amplicon analysis, such as HLA typing and all RNA quality checks.
ViiA 7 real-time PCR
The ViiA™ 7 Real-Time PCR System in our lab has the tube, 96-well and TLDA card accessories (along with approprioate centrifuge/rotors for loading TLDA cards). We mostly use TaqMan chemistry and 96-well FAST plates. The system offers exceptional reproducibility with minimal well-to-well (<3%) and plate-to-plate variation (<5%).
Quant Studio 12K flex
The QS12K offers a nanofluidics thermal cycler platform from Applied Biosystems™ offering the possibility to achieve highly reproducible and over 110,000 data points in a single eight-hour day. This instrument is currently available as part of the JDRF Australia microRNA biomarker project.
AccuFill Robotic System
The AccuFill™ system offers a fully automated robotic loading platform for OpenArray panels. It offers an easy-to-use software-driven automation integration and reduce hands on time. The AccuFill offers the ease of workflow for off-the-shelf as well as customized OA panels.
Robotic autodropet generator
The AutoDG offers the capacity to carry out automated partitioning into ~20,000 droplets/reaction for high throughput absolute transcript quantitation. The automated Droplet Generator platforms simplifies the ddPCR workflow, making digital PCR both scalable and practical. The AutoDG Instrument eliminates user-to-user variability with a more consistent droplet number (>16-18,000 droplets), allowing reproducibility between users as well as across laboratories (assessed between our lab and Raghu Mirmira's lab). The system generates droplets for 96 ddPCR reactions in under 45 minutes.
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QX-200 digital droplet (dd)PCR
The BioRad digital PCR system offers absolute quantitation of gene transcripts. This system has been used by the group in a JDRF Australia-funded project comp[aring circulating cfDNA in over 3000 samples. The system generally provides over 16,000 droplets for absolute quantitation.
QIAcuity One
The QIAcuity One (5-plex) Digital PCR System is designed to deliver precise and multiplexed quantification results for mutation detection, copy number variation (CNV), gene expression studies, gene-editing analysis, and many more. This nanoplate-based system seamlessly integrates a standard dPCR workflow of partitioning, thermocycling and imaging into a walk-away automated platform with minimal hands-on time.
Cell culture (Class II Type A2)
There are two cabinets that are designated for pre- and post-PCR workflows. These offer the capacity to carry all pre- and post-PCR workflows within the molecular biomarker laboratory.
Trace 1610 - Gas Chromatography Mass Spectrometry
The ThermoFisher Scientific Trace 1610 GCMS system is now commissioned and functional. The system is equipped with a Hydrogen and Nitrogen gas generator and has a Helium saver Hydrogen safe module. Different columns are available for untargeted metabolite analyses along with the Innowax column for the measurement of short chain fatty acids. The system has an autosampler and is also connected to a fraction collector (for seperate perifusion work) .
Qubit Flex Fluorometer
The Invitrogen Qubit Fluorometer is a popular benchtop fluorometer designed to accurately measure DNA, RNA, and protein quantity. Qubit is an easy-to-use touch screen menu driven system that make it easy to select and run the assays you need, with results displayed in just a few seconds. For more information, check: https://www.thermofisher.com/order/catalog/product/Q33327
Thermal cyclers (multiple systems)
There are multiple thermal cyclers in the main molecular lab. The BioRad thermal cycler is committed for the droplet digital PCR workflow while the Eppendorf and the Thermo (SimpliAmp) thermal cyclers are for routine reverse transcription and PCR workflows.
Islet stimulation studies
Islet perifusion system is now set up to allow perifusion of islet or islet-like clusters using a fraction collector that is linked to the Chromeleon software. The system would require some degree of training apart from the standard induction. PLease contact Prof. Hardikar for training and use of this system.
Cell culture (Class II Type A2)
There is a mouse/rat cell culture facility on level 1 and a human (primary cells/cell line) culture facility on the ground floor. There are inverted and fluorescent microscopy facilities in the rooms and automated cell counters for quick and easy detection of GFP/RFP-expressing cells. There are PC2 certified laboratory facilities with flow cytometry, Mass Spec, time-lapse imaging and fluorescent microscopy for visualizing lentiviral transformed cells. All of the puramycin selected and Dox-regulatable miRNA-lines should be used in these PC2 facilities following appropriate induction and training.
EVOS M7000
This autmicroscope offers brilliant images throughout the entire 8:1 zoom range and enables easy detection of developing pancreas buds (with the MIP-GFP / PDX1-GFP) reporter mice. The microscope is equipped with fluorescent filters to enable visualization of GFP.
Pre- and Post-PCR processing cabinets
Seperate pre- and post-PCR cabinets with their own sets of pipettes are placed in the main molecular biology lab on level 1. Do not remove the pipettes from these cabinets except for routine calibration and any decontamination, as required. Ensure that you turn on the UV before and after use.
Bioruptor
The Bioruptor® uses a unique system to uniformely process multiple samples in sealed 0.5 or 1.7ml eppendorf tubes. The built-in cooling system (water cooler and Single Cycle Valve) ensures high precision temperature control resulting in higher quality samples for shearing chromatin in chromatin immunoprecipitation (ChIP)-related workflows. The system can also be used for other applications associated with cell disruption.
Countess III automated cell counter
The Countess automated cell counter contain advanced autofocusing and counting algorithms to facilitate quickly and accurate counting of cells within a population. A standard (glass) haemocytometer slide is also compatible with the Countess III system. However, for fluorecent labeling counting, it is recommened to use the disposable slides. The system offers a significant advantage for automated cell counting, especially when working with multiple lines.
Surgeries
The Hardikar lab uses several animal models for relevant studies. These include high fat (up to 60% fat) or cafeteria diet (highly palatable diets) studies. The lab has expertise and track record over the past two decades to carry out surgeries including islet transplantation under the kidney capsule, via Theracytes or alginate-based microcapsules, panreatectomy, nephrectomy and delivery of specific compounds (eg. short chain fatty acids/SCFAs) to the gut using technologies that deliver continuous fatty acid and other supplements to specific regions of the gut. The lab also has experience and expertise in developing animal models of multigenerational undernutrition and central adiposity as well as models of high-fat-fed insulin resistance and Type 2 diabetes that help in assessing the potential of nutrient/pre-/pro-biotic supplements in diabetes. Contact Prof. Anand Hardikar for surgical assistance and/or animal experimentation.