ABSTRACT
We present the RespiDisk enabling the fully automated and multiplex point-of-care (POC) detection of (currently) up to 19 respiratory tract infection (RTI) pathogens from a single sample based on reverse transcriptase polymerase chain reaction (RT-PCR). RespiDisk comprises a RTI-specific implementation of the centrifugal microfluidic LabDisk platform and combines new and existing advanced unit operations for liquid control, thereby automating all assay steps only by a spinning frequency and temperature protocol in combination with the use of a permanent magnet for in situ bead handing. The capabilities of the system were demonstrated with 36 tested quality samples mimicking clinical conditions (clinical and/or cultured material suspended in transport medium or synthetic bronchoalveolar lavage (BAL)) from past external quality assessment (EQA) panels covering 13 of the 19 integrated RTI detection assays. In total, 36 samples × 19 assays/sample resulting in 684 assays were performed with the RespiDisk, and its analytical performance was in full agreement with the routine clinical workflow serving as reference. A strong feature of the platform is its universality since its components allow the simultaneous detection of a broad panel of bacteria and viruses in a single run, thereby enabling the differentiation between antibiotic-treatable diseases. Furthermore, the full integration of all necessary biochemical components enables a reduction of the hands-on time from manual to automated sample-to-answer analysis to about 5 min. The study was performed on an air-heated LabDisk Player instrument with a time-to-result of 200 min.
Subject(s)
Respiratory Tract Infections , Viruses , Bacteria , Humans , Microfluidics , Point-of-Care Systems , Respiratory Tract Infections/diagnosisABSTRACT
Two extremely bulky amidinate ligands, [RC{N(Dip)}{N(Ar)}]- (Dip = 2,6-diisopropylphenyl; Ar = C6H2{C(H)Ph2}2Pri-2,6,4; R = 1-adamantyl (LAd), tert-butyl (LtBu)) have been developed and utilised for the kinetic stabilisation of the strontium and magnesium hydride complexes, [LAdSr(µ-H)]2 and [LRMg(µ-H)]2 (R = Ad or But). The former represents the missing link in the series of dimeric systems, [LAe(µ-H)]2 (Ae = alkaline earth metal). The structure and bonding of the complexes have been studied by crystallographic, spectroscopic and computational techniques.
ABSTRACT
INTRODUCTION: Despite effective in vitro preclinical strategies to identify cardiovascular (CV) liabilities, there remains a need for early functional assessment prior to complex in vivo mammalian models. The larval zebrafish (Danio rerio, Zf) has been suggested for this role: previous data suggest that cardiac electrophysiology and vascular ultrastructure are comparable with mammals, and also indicate responsiveness of individual Zf CV system endpoints to some functional modulators. Little information is, however, available regarding integrated functional CV responses to drug treatment. Consequently, we developed a novel larval Zf model capable of simultaneous quantification of chronotropic, inotropic and arrhythmic effects, alongside measures of blood flow and vessel diameter. METHODS: Non-invasive video analysis of the heart and dorsal aorta of anaesthetized and agarose-embedded larval ZF was used to measure multiple cardiovascular endpoints, simultaneously, following treatment with a range of functional modulators of CV physiology. RESULTS: Changes in atrial and ventricular beat frequencies were detected in response to acute treatment with cardio-stimulants (adrenaline and theophylline), and negative chrono/inotropes (cisapride, haloperidol, terfenadine and verapamil). Arrhythmias were also observed including terfenadine-induced 2:1 atrial-ventricular (A-V) block, a previously proposed hERG surrogate measure. Significant increases in blood flow were detected in response to adrenaline and theophylline exposure; and decreases after cisapride, haloperidol, terfenadine, and verapamil treatment. Using dorsal aorta (DA) blood flow and ventricular beat rate, surrogate stoke volumes were also calculated for all compounds. DISCUSSION: These data support the use of this approach for CV function studies. Moreover the throughput and compound requirements (approximately 3 compounds/person effort/week and <10 mg) make our approach potentially suitable for higher throughput drug safety and efficacy applications, pending further assessment of ZF-mammalian pharmacological comparability.
