Aptamers are single-stranded oligonucleotide molecules that bind with high affinity and specificity to a wide range of target molecules. The method of systematic evolution of ligands by exponential enrichment (SELEX) plays an essential role in the isolation of aptamers from a randomized oligonucleotide library. To date, significant modifications and improvements of the SELEX process have been achieved, engendering various forms of SELEX from conventional SELEX to microfluidics-based full-chip SELEX. While full-chip SELEX is generally considered advantageous over conventional SELEX, there has not yet been a conclusive comparison between the methods. Herein, we present a comparative study of three SELEX strategies for aptamer isolation, including those using conventional agarose bead-based partitioning, microfluidic affinity selection, and fully integrated microfluidic affinity selection and PCR amplification. Using immunoglobulin E (IgE) as a model target molecule, we compare these strategies in terms of the time and cost for each step of the SELEX process including affinity selection, amplification, and oligonucleotide conditioning. Target-binding oligonucleotides in the enriched pools are sequenced and compared to assess the relative efficacy of the SELEX strategies. We show that the microfluidic strategies are more time- and cost-efficient than conventional SELEX.
Simultaneous infections of tuberculosis and toxoplasmosis are uncommon in immunocompetent patients. We report a case of a 30-year-old male who presented with right eye redness and blurring of vision for one month. The visual acuities were hand movement and Snellen 20/30 in the right and left eyes, respectively. Panuveitis and scleritis were found in the right eye, together with dense vitritis and a large choroidal lesion that extended from the inferonasal to the superotemporal quadrants. B-scan ultrasonography of the right eye showed a choroidal detachment with scleral thickening and subtenon fluid. His IgM anti-Toxoplasma antibody was detected, and his QuantiFERON Gold testing was positive. Magnetic resonance imaging (MRI) of the orbit revealed an enhancing intraocular mass at the lateral and inferior aspects of the right globe. The diagnosis of right eye sclerouveitis with presumed tuberculosis and toxoplasmosis co-infections was made. He was treated with a course of oral azithromycin and anti-tubercular therapy along with systemic prednisolone and a topical steroid. The treatment reduced the inflammation; however, the patient suffered from a sequela of chronic uveitis with prolonged hypotony. Medical treatment alone may be insufficient in treating severe infective sclerouveitis, hence surgical intervention might be warranted to provide favorable clinical outcomes.
The performance of aptamers as versatile tools in numerous analytical applications is critically dependent on their high target binding specificity and selectivity. However, only the technical or methodological aspects of measuring aptamer-target binding affinities are focused, ignoring the equally important mathematical components that play pivotal roles in affinity measurements. In this study, we aim to provide a comprehensive review regarding the utilization of different mathematical models and equations, along with a detailed description of the computational steps involved in mathematically deriving the binding affinity of aptamers against their specific target molecules. Mathematical models ranging from one-site binding to multiple aptameric binding site-based models are explained in detail. Models applied in several different approaches of affinity measurements such as thermodynamics and kinetic analysis, including cooperativity and competitive-assay based mathematical models have been elaborately discussed. Mathematical models incorporating factors that could potentially affect affinity measurements are also further scrutinized.