A multi-function, disposable, microfluidic module for mutation detection

Recognition of point mutations in a codon 12 of the K-ras gene, most frequently observed, is considered to be useful in the early diagnosis of several types of the human cancers. We have developed a multifunction, disposable, microfluidic module which detects low-abundant point mutations in human genomic DNA in modular architecture. Each functional component including a microfluidic PCR reactor, a passive diffusional micromixer reactor, and a microfluidic LDR reactor was separately designed and fabricated. Fluidic interconnects were also developed to make a fluidic passage between the functional components. Polycarbonate substrates were micro-molded, using hot embossing with micro-milled brass mold inserts to make all microfluidic components. Developed microassembly using passive alignment features, fabricated on all components, was used to assemble the functional components with the fluidic interconnects using an adhesive bonding technique. Thermal simulations were employed to ensure uniform thermal distributions in the microfluidic PCR and LDR reactors, to isolate the mixing junction in order to avoid heat–induced bubble formation in the passive micromixer reactor, and to have minimal thermal crosstalk due to the asymmetric thermal zones in the PCR and the LDR reactors. A control system was developed to control temperatures enabling thermal cycling in the microfluidic PCR and LDR reactor. LDR products were produced using the module within an hour with DNA sample, which had the ratio of 1:200. Total reaction time was about 67 minutes. By applying an enzyme as a purification of PCR products, a LDR analysis can be optimized and minimized to reduce the false positive signals and inconstant results generated by PCR products during the LDR. The purification system allowed us to successfully quantify the amount of mutant alleles in the genomic DNA. The high degree of accuracy in this module can also facilitate the detection of low-frequency point mutation occurred in other functional genes. This module, fabricated using replication technologies of polymers will be able to supply low cost, disposable detection tools for known disease-causing mutations and also expand to other PCR-based detection assays in diagnostic applications

Lactate: a multifunctional signaling molecule

Since its discovery in 1780, lactate has long been misunderstood as a waste by-product of anaerobic glycolysis with multiple deleterious effects. Owing to the lactate shuttle concept introduced in the early 1980s, a paradigm shift began to occur. Increasing evidence indicates that lactate is a coordinator of whole-body metabolism. Lactate is not only a readily accessible fuel that is shuttled throughout the body but also a metabolic buffer that bridges glycolysis and oxidative phosphorylation between cells and intracellular compartments. Lactate also acts as a multifunctional signaling molecule through receptors expressed in various cells and tissues, resulting in diverse biological consequences including decreased lipolysis, immune regulation, anti-inflammation, wound healing, and enhanced exercise performance in association with the gut microbiome. Furthermore, lactate contributes to epigenetic gene regulation by lactylating lysine residues of histones, accounting for its key role in immune modulation and maintenance of homeostasis

Closed-Form Solution of the Unit Normal Loss Integral in Two-Dimensions

In Value of Information (VoI) analysis, the unit normal loss integral (UNLI) frequently emerges as a solution for the computation of various VoI metrics. However, one limitation of the UNLI has been that its closed-form solution is available for only one dimension, and thus can be used for comparisons involving only two strategies (where it is applied to the scalar incremental net benefit). We derived a closed-form solution for the two-dimensional UNLI, enabling closed-form VoI calculations for three strategies. We verified the accuracy of this method via simulation studies. A case study based on a three-arm clinical trial was used as an example. VoI methods based on the closed-form solutions for the UNLI can now be extended to three-decision comparisons, taking a fraction of a second to compute and not being subject to Monte Carlo error. An R implementation of this method is provided as part of the predtools package (https://github.com/resplab/predtools/).Comment: 1 table, 1 figure, will be submitted to MDM - technical not