APEX protocol implementation on a lab-on-a-chip for SNPs detection

Abstract

In this work a detection module for the single nucleotide polymorphisms (SNPs) detection was realised. In particular arrayed primer extension (APEX) was selected as innovative method for SNPs detection and this protocol was scaled down following a micro total analysis approach in order to fabricate a lab-on-a-chip (LOC). Finite element analysis and behavioural simulations with commercial tools to properly design the microfluidic circuitry have preceded the technological processes for the production of the device. The fluidic was designed to contain 5μl of DNA and reagents that are inserted through three inlets. The layout includes two mixers and a sealed reaction chamber. Glass/silicon prototypes were fabricated with the employment of micro-electro-mechanical-system (MEMS) processes. The devices were tested with APEX biological protocols customized for the scaled volumes. Furthermore, in order to demonstrate the absence of any negative interaction between the chip and the APEX reagents, different chips were tested in intermediate steps together with the protocol executed in standard conditions. The final result demonstrates that the thermo sequenase extended the probes with the dideoxynucleotides modified with Cy5 fluorophore, thus indicating the possibility of implementation of the APEX protocol on LOC devices.