SARS-CoV-2 Aptasensors Based on Electrochemical Impedance Spectroscopy and Low-Cost Gold Electrode Substrates.

SARS-CoV-2 diagnostic practices broadly involve either quantitative polymerase chain reaction (qPCR)-based nucleic amplification of viral sequences or antigen-based tests such as lateral flow assays (LFAs). Reverse transcriptase-qPCR can detect viral RNA and is the gold standard for sensitivity. However, the technique is time-consuming and requires expensive laboratory infrastructure and trained staff. LFAs are lower in cost and near real time, and because they are antigen-based, they have the potential to provide a more accurate indication of a disease state. However, LFAs are reported to have low real-world sensitivity and in most cases are only qualitative. Here, an antigen-based electrochemical aptamer sensor is presented, which has the potential to address some of these shortfalls. An aptamer, raised to the SARS-CoV-2 spike protein, was immobilized on a low-cost gold-coated polyester substrate adapted from the blood glucose testing industry. Clinically relevant detection levels for SARS-CoV-2 are achieved in a simple, label-free measurement format using sample incubation times as short as 15 min on nasopharyngeal swab samples. This assay can readily be optimized for mass manufacture and is compatible with a low-cost meter.

Pharmacokinetics and pharmacodynamics of a high concentration of buprenorphine (Simbadol) in conscious horses after subcutaneous administration.

OBJECTIVE: To determine the pharmacokinetics and pharmacodynamics of high-concentration formulation of buprenorphine (1.8 mg mL-1; Simbadol) following subcutaneous (SC) administration in horses. STUDY DESIGN: Prospective, randomized, crossover trial. ANIMALS: A group of six healthy adult horses weighing 521-602 kg. METHODS: On three occasions, Simbadol (0.005 mg kg-1; treatment S5), (0.0025 mg kg-1; treatment S2.5) or saline (treatment SAL) were administered SC at least 7 days apart in random order. Electrical nociceptive threshold (ENT) measured on the neck region, physiologic variables, locomotor activity, degree of restlessness and presence of excitatory signs were measured at baseline and for up to 48 hours after injection. Blood was collected for pharmacokinetic analysis at the same time intervals and plasma buprenorphine concentration (Cp) measured using liquid chromatography-tandem mass spectrometry. RESULTS: Buprenorphine was quantifiable in all horses from 15 minutes after administration up to 8-12 hours. ENT was significantly increased in treatment S2.5 compared with treatment SAL at 0.75-6 hours after treatment. Increase in locomotor activity and compulsive behavior were recorded in all horses after Simbadol, and degree of restlessness was significantly higher in treatment S5 than SAL for a sustained time. Gastrointestinal motility significantly decreased in all horses after Simbadol and returned to baseline by 16 hours after treatment. CONCLUSIONS AND CLINICAL RELEVANCE: In horses, SC Simbadol was rapidly absorbed and Cp decreased rapidly. Side effects commonly seen in horses after opioids were observed in both Simbadol treatments, but degree of opioid-induced excitement lasted significantly longer in treatment S5. Simbadol (0.0025 mg kg-1) SC has the potential to be used clinically to treat pain in horses. However, at this dose, duration of antinociceptive effects was not longer than that reported for conventional buprenorphine, and side effects, including reduction in gastrointestinal motility and increased locomotor activity, were documented.

Morphine synovial fluid concentrations after intravenous regional limb perfusion in horses during standing sedation.

BACKGROUND: Addition of morphine to the perfusate while performing intravenous regional limb perfusion (IVRLP) may be helpful in treating painful infectious orthopaedic conditions of the distal limb. OBJECTIVES: The main objective of this study was to determine synovial morphine concentrations following IVRLP with morphine alone or in combination with amikacin. STUDY DESIGN: Randomised cross-over in vivo experiment. METHODS: Six horses underwent IVRLP with 0.1 mg/kg morphine sulphate diluted to 60 mL using 0.9% NaCl (M group) or combined with 2 g amikacin and 0.9% NaCl (MA group) with a 2-week washout period between treatments. Synovial fluid was collected from the radiocarpal joint (RCJ) at 10, 20, 30, 120, 240, 480, 720 and 1440 min after IVRLP. The tourniquet was removed after the 30-min sample was collected. Synovial concentrations of morphine and major metabolites were measured using liquid chromatography-tandem mass spectrometry. Amikacin concentrations were quantified by a fluorescence polarisation immunoassay. RESULTS: Measurable concentrations of morphine were apparent in the RCJ of all horses. Median CMAX of morphine in the M group was 4753.1 (2115.7-14 934.5) ng/mL and 4477 (3434.3-7363) ng/mL in the MA group (p = 0.5). Median CMAX of synovial amikacin was 322.6 (157.5-1371.6 µg/mL). MAIN LIMITATIONS: Limitations include small sample size. Investigators were not blinded to the treatments and a third treatment group where amikacin alone was administered via IVRLP to the study population was not included. CONCLUSIONS: IVRLP using morphine is a feasible technique and synovial morphine concentrations were measurable following IVRLP and were not affected when used concurrently with amikacin. Administration of morphine via IVRLP may be beneficial as an analgesic technique for orthopaedic conditions of the distal limb while limiting potential serious systemic side-effects.

Treatment of lidocaine-refractory ventricular tachycardia using procainamide in an anesthetized horse.

OBJECTIVE: To highlight the use of procainamide as a potential alternative treatment modality in cases of ventricular tachycardia that are refractory to lidocaine and magnesium sulfate. ANIMAL: 1 adult horse weighing 380 kg. CLINICAL PRESENTATION, PROGRESSION, AND PROCEDURES: A 25-year-old Arabian gelding presented with severe colic signs. Due to persistent pain, it was elected to carry out an exploratory laparotomy. During the procedure a diagnosis of severe, unstable ventricular tachycardia was made based on the ECG findings, with an initial heart rate of 195 beats per minute and severe hypotension. TREATMENT AND OUTCOME: Initial treatment consisted of discontinuing dobutamine and the administration of a 2 mg/kg IV lidocaine bolus followed by a continuous rate infusion at 50 µg/kg/min. Twenty grams magnesium sulfate (5 mg/kg) was administered IV in 1 L of lactated Ringer solution as a slow bolus over 30 minutes. Ventricular tachycardia persisted with poor peripheral pulses, a severely dampened arterial waveform, and a MAP of 30 to 45 mm Hg. Two milligrams/kg IV procainamide was administered over 3 minutes, 3 separate times, at 5-minute intervals. Immediately following the third dose sinus rhythm was detected on the ECG, the arterial waveform improved, and MAP increased to 85 mm Hg. CLINICAL RELEVANCE: Ventricular tachycardia is a rare but potentially life-threatening complication in horses undergoing general anesthesia. The potential of this arrhythmia to progress to ventricular fibrillation is of grave concern, as the option to attempt to externally defibrillate horses back to normal sinus rhythm does not exist. This case highlights procainamide as a potential option for cases of ventricular tachycardia that are refractory to more standard treatment modalities.

Highly Selective Aptamer-Molecularly Imprinted Polymer Hybrids for Recognition of SARS-CoV-2 Spike Protein Variants.

Virus recognition has been driven to the forefront of molecular recognition research due to the COVID-19 pandemic. Development of highly sensitive recognition elements, both natural and synthetic is critical to facing such a global issue. However, as viruses mutate, it is possible for their recognition to wane through changes in the target substrate, which can lead to detection avoidance and increased false negatives. Likewise, the ability to detect specific variants is of great interest for clinical analysis of all viruses. Here, a hybrid aptamer-molecularly imprinted polymer (aptaMIP), that maintains selective recognition for the spike protein template across various mutations, while improving performance over individual aptamer or MIP components (which themselves demonstrate excellent performance). The aptaMIP exhibits an equilibrium dissociation constant of 1.61 nM toward its template which matches or exceeds published examples of imprinting of the spike protein. The work here demonstrates that "fixing" the aptamer within a polymeric scaffold increases its capability to selectivity recognize its original target and points toward a methodology that will allow variant selective molecular recognition with exceptional affinity.