Digital microfluidic platform for the detection of rubella infection and immunity: a proof of concept.

BACKGROUND: Whereas disease surveillance for infectious diseases such as rubella is important, it is critical to identify pregnant women at risk of passing rubella to their offspring, which can be fatal and can result in congenital rubella syndrome (CRS). The traditional centralized model for diagnosing rubella is cost-prohibitive in resource-limited settings, representing a major obstacle to the prevention of CRS. As a step toward decentralized diagnostic systems, we developed a proof-of-concept digital microfluidic (DMF) diagnostic platform that possesses the flexibility and performance of automated immunoassay platforms used in central facilities, but with a form factor the size of a shoebox. METHODS: DMF immunoassays were developed with integrated sample preparation for the detection of rubella virus (RV) IgG and IgM. The performance (sensitivity and specificity) of the assays was evaluated with serum and plasma samples from a commercial antirubella mixed-titer performance panel. RESULTS: The new platform performed the essential processing steps, including sample aliquoting for 4 parallel assays, sample dilution, and IgG blocking. Testing of performance panel samples yielded diagnostic sensitivity and specificity of 100% and 100% for both RV IgG and RV IgM. With 1.8 µL sample per assay, 4 parallel assays were performed in approximately 30 min with <10% mean CV. CONCLUSIONS: This proof of concept establishes DMF-powered immunoassays as being potentially useful for the diagnosis of infectious disease.

Automated digital microfluidic platform for magnetic-particle-based immunoassays with optimization by design of experiments.

We introduce an automated digital microfluidic (DMF) platform capable of performing immunoassays from sample to analysis with minimal manual intervention. This platform features (a) a 90 Pogo pin interface for digital microfluidic control, (b) an integrated (and motorized) photomultiplier tube for chemiluminescent detection, and (c) a magnetic lens assembly which focuses magnetic fields into a narrow region on the surface of the DMF device, facilitating up to eight simultaneous digital microfluidic magnetic separations. The new platform was used to implement a three-level full factorial design of experiments (DOE) optimization for thyroid-stimulating hormone immunoassays, varying (1) the analyte concentration, (2) the sample incubation time, and (3) the sample volume, resulting in an optimized protocol that reduced the detection limit and sample incubation time by up to 5-fold and 2-fold, respectively, relative to those from previous work. To our knowledge, this is the first report of a DOE optimization for immunoassays in a microfluidic system of any format. We propose that this new platform paves the way for a benchtop tool that is useful for implementing immunoassays in near-patient settings, including community hospitals, physicians' offices, and small clinical laboratories.

Comparison of the accuracy of radiography and ultrasonography for detection of articular lesions in horses.

OBJECTIVE: To compare the accuracy of ultrasonographic and radiographic examination for evaluation of articular lesions in horses. STUDY DESIGN: Cross-sectional study. ANIMALS: Horses (n = 137) with articular lesions. METHODS: Radiographic and ultrasonographic examinations of the affected joint(s) were performed before diagnostic or therapeutic arthroscopic surgery. Findings were recorded and compared to lesions identified during arthroscopy. RESULTS: In 254 joints, 432 lesions were identified by arthroscopy. The overall accuracy was 82.9% for ultrasonography and 62.2% for radiography (P < .0001) with a sensitivity of 91.4% for ultrasonography and 66.7% for radiography (P < .0001). The difference in specificity was not statistically significant (P = .2628). The negative predictive value for ultrasonography was 31.5% and 13.2% for radiography (P = .0022), the difference for the positive predictive value was not statistically significant (P = .3898). The accuracy for ultrasonography and radiography for left versus right joints was equal and corresponded with the overall results. CONCLUSIONS: Ultrasonographic evaluation of articular lesions was more accurate than radiographic evaluation.

Removal of equine cryptorchid testes through an enlarged umbilical portal in dorsally recumbent horses after intra-abdominal laparoscopic castration.

BACKGROUND: Laparoscopic removal of cryptorchid testes has been routinely reported through enlarged parainguinal incisions in dorsally recumbent horses. Outcomes following removal through an extended umbilical incision have not been previously reported. OBJECTIVE: To describe the surgical technique of removing cryptorchid testes in dorsally recumbent horses through an enlarged umbilical portal after laparoscopic intra-abdominal castration. STUDY DESIGN: Retrospective case series. METHODS: Medical records were reviewed for horses that underwent laparoscopic removal of unilateral or bilateral cryptorchid testes from January 2006 to December 2016. Horses were placed under general anaesthesia, positioned in dorsal recumbency and then tilted into Trendelenburg position. Cryptorchid testes were castrated by ligating loop application and/or electrosurgery. The umbilical portal incision was extended along the linea alba for testes removal. All descended testes were removed by routine closed castration with the scrotal incision left to heal by second intention. Perianaesthetic laboratory values, surgical procedure descriptions, surgery and anaesthesia times, and in-hospital perioperative complications were recorded. RESULTS: A total of 79 horses, aged 1-14 years, with unilateral or bilateral cryptorchidism were included: 90 cryptorchid testes were successfully removed through an enlarged umbilical portal incision. Sixty-eight horses were unilaterally cryptorchid and 11 were bilaterally cryptorchid. Two horses had minor post-operative complications related to the extended umbilical portal incision. MAIN LIMITATIONS: Retrospective nature of the study, no control population for comparison and no follow-up after hospital discharge. CONCLUSION: An extended umbilical portal incision is a successful alternative to extending a parainguinal incision for testis removal after laparoscopic castration.

Postoperative complications and survival after enterolith removal from the ascending or descending colon in horses.

OBJECTIVE: To compare incidence of postoperative complications and survival of horses that had surgery for enterolithiasis of the ascending (AC) or descending (DC) colon. STUDY DESIGN: Case series ANIMALS: Horses (n=236) that had celiotomy for AC (n=97) or DC (n=139) enterolithiasis. METHODS: Medical records (1999-2005) were reviewed for signalment, presenting clinical signs, surgical findings, postoperative complications, and short-term survival until discharge. Information on performance and survival > or = 1 year was obtained by telephone or mailed questionnaire. RESULTS: Number of postoperative complications did not differ between groups (P=.76). The most frequently identified short-term complications for all horses were incisional problems, gastric ulceration, ileus, diarrhea, fever, and anorexia. Horses with DC enteroliths had a significantly higher incidence of anorexia (P=.04) and fever (P=.01). The most common complications after hospital discharge were incisional problems, laminitis, weight loss, and colic. Although more DC horses were euthanatized intraoperatively (P=.02), no differences were detected for horses that survived until discharge (P=.18) or > or = 1 year after discharge (P=.47). CONCLUSIONS: Number or type of postsurgical complications or survival after surgery was not influenced by enterolith location and horses have a favorable prognosis for long-term survival after enterolith removal, regardless of site of obstruction in the colon. CLINICAL RELEVANCE: Incidence of postoperative complications and survival are not affected by the site of enterolith obstruction in the colon.

A study of the liquid and solvent properties of optically active and racemic alpha-methylbenzylamine.

Speed-of-sound measurements are reported for RS (racemic) and S liquid alpha-methylbenzylamine (MBA) obtained using a modified design of previously published experimental geometry. After correcting for density changes, the resulting isentropic compressibility of the S liquid is found to be 2% larger than that of the RS racemic mixture. These data, along with proton NMR chemical shifts and published partial molar volumes, suggest that the structures of the racemic and optically active liquids are subtly different. The magnitude of the compressibilities and other data such as viscosity and the Kamlet-Taft parameters are consistent with the molecules in the liquids interacting via extensive hydrogen bonding. Cyclohexane, toluene, nitrobenzene, dimethyl sulfoxide, and methanol are completely miscible in MBA, as predicted from their corresponding Hildebrand solubility parameters. Proton NMR, optical rotation, and IR studies were carried out on solutions of the five solutes in MBA as a function of mole fraction. The asymmetric NH stretch of MBA was particularly informative in this regard. Solvation models were developed for the five solutes. The NH stretch frequency in cyclohexane changed only slightly up to a mole fraction of 0.7 in the hydrocarbon at which point the NH stretch greatly increased by about 200 cm(-1) at a mole fraction of 1. The hydrogen-bonding network does not dissipate until there is at least 70 mol % cyclohexane in the MBA. Gaussian calculations were also carried out to complement the ir studies.

Solvent effects on the optical rotation of (S)-(-)-alpha-methylbenzylamine.

The optical rotation of (S)-(-)-alpha-methylbenzylamine at 589 nm has been measured in 39 different solvents at five different concentrations: 0.25, 0.50, 1.00, 2.00, and 3.00 M. A correlation of the intrinsic rotations (i.e., extrapolation of specific rotations to zero concentration) with Kamlet's and Taft's solvent parameters (alpha, beta, and pi) is established. The polarity/polarizability, pi, and solvent acidity, alpha, terms are found to have a greater effect upon the optical rotation than the basicity of the solvent, beta. The specific rotation for (S)-(-)-alpha-methylbenzylamine has been calculated with Gaussian03 using a PCM model (B3LYP aug-cc-pVDZ) for all 39 solvated systems. Comparisons between the experimental and calculated values show the importance of hydrogen bonding on specific rotation.