A simple, accurate and cost-effective capillary electrophoresis test with computational methods to aid in universal microsatellite instability testing

Background: Microsatellite instability (MSI) testing is important for the classification of Lynch syndrome, as a prognostic marker and as a guide for adjuvant chemotherapy in colorectal cancer (CRC). The gold standard for determining MSI status has traditionally been fluorescent multiplex polymerase chain reaction (PCR) and capillary gel electrophoresis (CGE). However, its use in the clinical setting has diminished and has been replaced by immunohistochemical (IHC) detection of loss of mismatch repair protein expression due to practicability and cost. The aim of this study was to develop a simple, cost-effective and accurate MSI assay based on CGE. Method: After amplification of microsatellites by polymerase chain reaction (PCR) using the National Cancer Institute (NCI) panel (BAT 25, BAT26, D5S346, D2S123, D17S250) of MSI markers, parallel CGE was utilized to classify colorectal cancers as MSI-H, MSI-L and MSS using the 5200 Fragment Analyzer System. Cell lines and patient cancer specimens were tested. DNA from 56 formalin-fixed paraffin-embedded cancer specimens and matched normal tissue were extracted and CGE was performed. An automated computational algorithm for MSI status determination was also developed. Results: Using the fragment analyser, MSI status was found to be 100% concordant with the known MSI status of cell lines and was 86% and 87% concordant with immunohistochemistry (IHC) from patient cancer specimens using traditional assessment and our MSI scoring system, respectively, for MSI determination. The misclassification rate was mainly attributed to IHC, with only one (1.8%) sampling error attributed to CGE testing. CGE was also able to distinguish MSI-L from MSI-H and MSS, which is not possible with IHC. An MSI score based on total allelic variability that can accurately determine MSI status was also successfully developed. A significant reduction in cost compared with traditional fluorescent multiplex PCR and CGE was achieved with this technique. Conclusions: A simple, cost-effective and reliable method of determining MSI status and an MSI scoring system based on an automatic computational algorithm to determine MSI status, as well as degree of allelic instability in colorectal cancer, has been developed using the 5200 Fragment Analyzer System

Undoped and Nickel-Doped Zinc Oxide Thin Films Deposited by Dip Coating and Ultrasonic Spray Pyrolysis Methods for Propane and Carbon Monoxide Sensing Applications

Undoped and nickel-doped zinc oxide thin films were deposited on sodalime glass substrates by utilizing dip coating and ultrasonic spray pyrolysis deposition techniques. In both cases zinc acetate and nickel acetylacetonate were used as zinc precursor and nickel dopant source, respectively. XRD analysis confirms the ZnO wurtzite structure with (002) as the preferential orientation.SEM studies show the formation of two types of morphologies, primarily a porous spherical grains with a grain size distribution from 40 to 150 nm and another, rose-like structures with size distribution from 30 to 200 nm, based on different deposition techniques utilized. The elemental depth profiles across the films were investigated by the secondary-ion mass spectrometry (SIMS). Different gas sensing responses of all ZnO films were obtained for both propane and carbon monoxide gases, at different gas concentrations and operating temperatures. The highest sensing response (~6) for undoped ZnO films was obtained for films deposited by ultrasonic spray pyrolysis (USP). Nevertheless, the highest sensing response (~4 × 104) for doped ZnO films was obtained for films deposited by dip coating method. The behavior of sensing responses is explained in detail based on the morphological properties and the amount of Ni impurities incorporated into the crystal lattice

A Study of the CO Sensing Responses of Cu-, Pt- and Pd-Activated SnO2 Sensors: Effect of Precipitation Agents, Dopants and Doping Methods

In this work, we report the synthesis of Cu, Pt and Pd doped SnO2 powders and a comparative study of their CO gas sensing performance. Dopants were incorporated into SnO2 nanostructures using chemical and impregnation methods by using urea and ammonia as precipitation agents. The synthesized samples were characterized using X-ray diffraction (XRD), Raman spectroscopy, scanning electron microscopy (SEM) and high resolution transmission electron microscopy (HR-TEM). The presence of dopants within the SnO2 nanostructures was evidenced from the HR-TEM results. Powders doped utilizing chemical methods with urea as precipitation agent presented higher sensing responses compared to the other forms, which is due to the formation of uniform and homogeneous particles resulting from the temperature-assisted synthesis. The particle sizes of doped SnO2 nanostructures were in the range of 40–100 nm. An enhanced sensing response around 1783 was achieved with Cu-doped SnO2 when compared with two other dopants i.e., Pt (1200) and Pd:SnO2 (502). The high sensing response of Cu:SnO2 is due to formation of CuO and its excellent association and dissociation with adsorbed atmospheric oxygen in the presence of CO at the sensor operation temperature, which results in high conductance. Cu:SnO2 may thus be an alternative and cost effective sensor for industrial applications

A Simple, Accurate and Cost-Effective Capillary Electrophoresis Test with Computational Methods to Aid in Universal Microsatellite Instability Testing

Background: Microsatellite instability (MSI) testing is important for the classification of Lynch syndrome, as a prognostic marker and as a guide for adjuvant chemotherapy in colorectal cancer (CRC). The gold standard for determining MSI status has traditionally been fluorescent multiplex polymerase chain reaction (PCR) and capillary gel electrophoresis (CGE). However, its use in the clinical setting has diminished and has been replaced by immunohistochemical (IHC) detection of loss of mismatch repair protein expression due to practicability and cost. The aim of this study was to develop a simple, cost-effective and accurate MSI assay based on CGE. Method: After amplification of microsatellites by polymerase chain reaction (PCR) using the National Cancer Institute (NCI) panel (BAT 25, BAT26, D5S346, D2S123, D17S250) of MSI markers, parallel CGE was utilized to classify colorectal cancers as MSI-H, MSI-L and MSS using the 5200 Fragment Analyzer System. Cell lines and patient cancer specimens were tested. DNA from 56 formalin-fixed paraffin-embedded cancer specimens and matched normal tissue were extracted and CGE was performed. An automated computational algorithm for MSI status determination was also developed. Results: Using the fragment analyser, MSI status was found to be 100% concordant with the known MSI status of cell lines and was 86% and 87% concordant with immunohistochemistry (IHC) from patient cancer specimens using traditional assessment and our MSI scoring system, respectively, for MSI determination. The misclassification rate was mainly attributed to IHC, with only one (1.8%) sampling error attributed to CGE testing. CGE was also able to distinguish MSI-L from MSI-H and MSS, which is not possible with IHC. An MSI score based on total allelic variability that can accurately determine MSI status was also successfully developed. A significant reduction in cost compared with traditional fluorescent multiplex PCR and CGE was achieved with this technique. Conclusions: A simple, cost-effective and reliable method of determining MSI status and an MSI scoring system based on an automatic computational algorithm to determine MSI status, as well as degree of allelic instability in colorectal cancer, has been developed using the 5200 Fragment Analyzer System