A Lab-on-a-Chip Based Automatic Platform for Continuous Nitrites Sensing in Aquaculture

In aquaculture, the density of fish stock, use of feeding, and surrounding environmental conditions can easily result in an excessive concentration of harmful compounds that require continuous monitoring. Chemical sensors are available for most of these compounds, however, operative conditions and continuous monitoring in water make the development of sensors suitable for long and unattended deployments difficult. A possible solution is the development of engineered automatic labs where the uptake of sample and the contact with water is reduced and the use of a minimal quantity of reagents enables the implementation of reliable chemical assays. In this paper, a platform for automatic chemical assays is presented. The concept is demonstrated with the detection of nitrites based on the well-known colorimetric Griess reaction. The platform is centered around a lab-on-a-chip where reagents and water samples are mixed. The color of the reaction product is measured with low-cost optoelectronic components. Results show the feasibility of the approach with a minimum detectable concentration of about 0.1 mg/L which is below the tolerance level for aquaculture farms.

Determination of Postmortem Interval by Estimating CSF Proteins Concentration after Death, by Dye Binding Method at a Tertiary Care Hospital in Lahore, Pakistan

Introduction: Postmortem interval (PMI) is the time lapse between death of a person and its postmortem examination i.e. autopsy. Estimating the postmortem interval (PMI) is an imperative perspective of forensic medicine. Aims & Objectives: This study was conducted to see the impact of CSF protein estimation on determination of PMI. Place and duration of study: It was an observational correlational study, conducted for one year at Department of Forensic Medicine, King Edward Medical University Lahore. Material & Methods: A total of 119 cadavers were included in this study. Chemicals used were disodium molybdate, pyrogallol, succinic acid. The minimum detectable concentration of total proteins in CSF with dye binding method using pyrogallol red was determined as 0.022g/l. Two ml of CSF was taken from each cadaver. Clear, colorless samples were taken in test tubes. Turbid and blood contained samples were not included in study. Protein concentration was determined using photospectrometry. Statistical analysis was done by SPSS-23. Quantitative variables like age were presented as mean ± SD. Qualitative variables like gender were presented as frequency and percentages. For comparison between PMI and CSF proteins concentration correlation was applied. Results: Males accounted for majority of our subjects. The mean value of proteins in CSF was 219.91± 113.121 mg/dl. The most common PMI was 11 to 20 hours. CSF proteins increased gradually over 72 hours after death. The results of present study showed a significant positive correlation between time of death and CSF proteins concentration after death. Conclusion: It is observed that as a whole protein concentration increased with increasing time of death. Hence, CSF protein concentration can be used in estimating time since death.

Obtaining and Characterization of Hybridomas Producing Monoclonal Antibodies to Shiga-Like Toxins of I and II Types

Objective – obtaining and characterization of hybrid cell lines producing monoclonal antibodies against I and II types of shiga-like toxins.Materials and methods. Shiga-like toxins obtained in “48thCentral Research Institute” of Ministry of Defense of Russian Federation (Kirov), BALB/c mice, myeloma cells SP2/0-Ag14 were used in research. Immune splenocytes and SP2/0-Ag14 myeloma cells were fused according to G. Kohler and C. Milstein method in De St. Fazekas and D. Scheidegger modifcation using 50 % polyethylene glycol. Hybrid cell lines producing specifc monoclonal antibodies were cloned by limited dilutions. Hybridomas growth and producing properties were studied in vitro and in vivo. Specifc activity of immune sera, culture and ascitic fluids were studied by indirect ELISA. Monoclonal antibodies from ascitic fluids were precipitated by saturated ammonium sulfate, followed by ion exchange chromatographyResults and discussion. 8 hybridomas producing monoclonal antibodies against I and II types shiga-like toxins were obtained. Hybridomas are characterized by stable proliferation and antibody-producing activity during 10 passages in vitro and 3 passages in vivo (observation period). Obtained monoclonal antibodies can be used for ELISA detection of I and II types shiga-like toxins. Minimum detectable concentration of shiga-like toxins in sandwich ELISA is 1 ng/ml. The possibility of detecting shiga-like toxins without typical differentiation was shown when using in the enzyme immunoassay a polyreceptor mixture of monoclonal antibodies for sensitizing the plate and a polyspecifc mixture of immunoperoxidase conjugates.

Millimeter-Wave-Based Spoof Localized Surface Plasmonic Resonator for Sensing Glucose Concentration

Glucose-monitoring sensors are necessary and have been extensively studied to prevent and control health problems caused by diabetes. Spoof localized surface plasmon (LSP) resonance sensors have been investigated for chemical sensing and biosensing. A spoof LSP has similar characteristics to an LSP in the microwave or terahertz frequency range but with certain advantages, such as a high-quality factor and improved sensitivity. In general, microwave spoof LSP resonator-based glucose sensors have been studied. In this study, a millimeter-wave-based spoof surface plasmonic resonator sensor is designed to measure glucose concentrations. The millimeter-wave-based sensor has a smaller chip size and higher sensitivity than microwave-frequency sensors. Therefore, the microfluidic channel was designed to be reusable and able to operate with a small sample volume. For alignment, a polydimethylsiloxane channel was simultaneously fabricated using a multilayer bonding film to attach the upper side of the pattern, which is concentrated in the electromagnetic field. This real-time sensor detects the glucose concentration via changes in the S11 parameter and operates at 28 GHz with an average sensitivity of 0.015669 dB/(mg/dL) within the 0–300 mg/dL range. The minimum detectable concentration and the distinguishable signal are 1 mg/dL and 0.015669 dB, respectively, from a 3.4 μL sample. The reusability and reproducibility were assessed through replicates.

CRISPR/Cas12a-Based Ultrasensitive and Rapid Detection of JAK2 V617F Somatic Mutation in Myeloproliferative Neoplasms

The JAK2 V617F mutation is a major diagnostic, therapeutic, and monitoring molecular target of Philadelphia-negative myeloproliferative neoplasms (MPNs). To date, numerous methods of detecting the JAK2 V617F mutation have been reported, but there is no gold-standard diagnostic method for clinical applications. Here, we developed and validated an efficient Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)/CRISPR associated protein 12a (Cas12a)-based assay to detect the JAK2 V617F mutation. Our results showed that the sensitivity of the JAK2 V617F/Cas12a fluorescence detection system was as high as 0.01%, and the JAK2 V617F/Cas12a lateral flow strip assay could unambiguously detect as low as 0.5% of the JAK2 V617F mutation, which was much higher than the sensitivity required for clinical application. The minimum detectable concentration of genomic DNA achieved was 0.01 ng/μL (~5 aM, ~3 copies/μL). In addition, the whole process only took about 1.5 h, and the cost of an individual test was much lower than that of the current assays. Thus, our methods can be applied to detect the JAK2 V617F mutation, and they are highly sensitive, rapid, cost-effective, and convenient.

Cas12a and Lateral Flow Strip-Based Test for Rapid and Ultrasensitive Detection of Spinal Muscular Atrophy

Spinal muscular atrophy (SMA) is characterized by severe lethality and irreversible progression. Early diagnosis of SMA is of more practical significance with the emergence of effective therapy. However, existing techniques to identify SMA patients rely on cumbersome instruments, hindering their accessibility and application. An SMA-Cas12a-strip assay was developed with the integration of Cas12a-based nucleic acid detection, isothermal amplification, and lateral flow strip. The analytical performance of the assay was assessed with clinical samples. To explore its extensible utility, various specimens were tested. Validated with 168 clinical samples, the sensitivity and specificity of the SMA-Cas12a-strip assay were both 100%. The minimum detectable concentration of genomic DNA containing the target gene achieved 526 aM. The assay was compatible with specimens from several sources, and the turnaround time could be within 1.5 h. We developed a simple, cost-effective, and highly sensitive and specific assay to detect SMA patients. With little and field-portable equipment, the assay holds great promise in the detection of SMA patients, particularly in low-resource regions.

Fabrication of Nano/Micro-Structured Electrospun Detection Card for the Detection of Pesticide Residues

A novel nano/micro-structured pesticide detection card was developed by combining electrospinning and hydrophilic modification, and its feasibility for detecting different pesticides was investigated. Here, the plain and hydrophilic-modified poly(ε-caprolactone) (PCL) fiber mats were used for the absorption of indolyl acetate and acetylcholinesterase (AChE), respectively. By pre-treating the fiber mat with ethanol, its surface wettability was improved, thus, promoting the hydrolysis of the PCL fiber mat. Furthermore, the absorption efficiency of AChE was improved by almost two times due to the increased hydrophilicity of the modified fiber mat. Noteworthily, this self-made detection card showed a 5-fold, 2-fold, and 1.5-fold reduction of the minimum detectable concentration for carbofuran, malathion, and trichlorfon, respectively, compared to the national standard values. Additionally, it also exhibited good stability when stored at 4 °C and room temperature. The food detection test showed that this nano/micro-based detection card had better detectability than the commercial detection card. Therefore, this study offers new insights into the design of pesticide detection cards, which also broadens the application of electrospinning technique.

CAESIUM RETENTION CHARACTERISTICS OF KNIFC–PAN RESIN FROM RIVER WATER

The caesium retention characteristics of a potassium-nickel hexacyanoferrate resin in a polyacrylnitrile (KNiFC–PAN) matrix were tested in fresh water over the range of 2.5–400 mL min−1. The experimental setup used 2 mL resin and 4-L aliquots of freshwater samples. The results showed nearly 100% retention at speeds below 10 mL min−1, above 80% up to 100 mL min−1, and approached 50% at 400 mL min−1. Using 100 mL min−1 flow rate and KNiFC–PAN resin in a well-type HPGe detector, the minimum detectable concentration was reduced to 3 mBq kg−1 for 4-L aliquots of water samples from the previous 15 mBq kg−1 achieved by Powdex ion-exchange resin and a planar type HPGe detector.

Real-Time Noninvasive Measurement of Glucose Concentration Using a Modified Hilbert Shaped Microwave Sensor

We developed a microwave glucose sensor based on the modified first-order Hilbert curve design and measured glucose concentration in aqueous solutions by using a real-time microwave near-field electromagnetic interaction technique. We observed S21 transmission parameters of the sensor at resonant frequencies depend on the glucose concentration. We could determine the glucose concentration in the 0–250 mg/dL concentration range at an operating frequency of near 6 GHz. The measured minimum detectable signal was 0.0156 dB/(mg/dL) and the measured minimum detectable concentration was 1.92 mg/dL. The simulation result for the minimum detectable signal and the minimum detectable concentration was 0.0182 dB/(mg/dL) and 1.65 mg/dL, respectively. The temperature instability of the sensor for human glycemia in situ measurement range (27–34 °C for fingers and 36–40 °C for body temperature ranges) can be improved by the integration of the temperature sensor in the microwave stripline platform and the obtained data can be corrected during signal processing. The microwave signal–temperature dependence is almost linear with the same slope for a glucose concentration range of 50–150 mg/dL. The temperature correlation coefficient is 0.05 dB/°C and 0.15 dB/°C in 27–34 °C and 36–40 °C temperature range, respectively. The presented system has a cheap, easy fabrication process and has great potential for non-invasive glucose monitoring.