Genotyping from saliva with a one-step microdevice

Abstract

This paper presents a disposable microfluidic device for on-chip lysing, PCR, and analysis in one continuous-flow process. Male-female sex determination was performed with human saliva in less than 20 min from spit to finish, and requiring only seconds of manual sample handling. This genetic analysis was based on the amplification and detection of the DYZ1 repeat region unique to the Y-chromosome. The flow-through microfluidic chip consisted of a single serpentine channel designed to guide samples through 42 heating and cooling cycles. Cycling was performed by matching the local channel geometry to a steady-state temperature gradient established across the microfluidic chip. 38 channel segments were designed for rapid low volume PCR, and four were optimized for spatial DNA melting analysis. Fluorescence detection was used to monitor the amplification and to capture the melting signature of the amplicon was performed with a basic 8-bit CCD camera. The microfluidic device itself was fabricated from microscope slides and a double-sided tape. The simplicity of the system and its robust performance combine in an elegant solution for lab-on-a-chip genetic analysis.

A study on the limits and advantages of using a desktop cutter plotter to fabricate microfluidic networks

“…Leaching studies have not been performed by us or by the manufacturer of this particular adhesive and may be necessary for some applications. However, a number of PSA films have been used for PCR, DNA and electrophoresis applications with good results, for example those by Nath et al (2014), Pjescic and Crews (2012) and Greer et al (2007). Solvents do attack both the adhesive film and a number of plastics.…”

“…The term was introduced in 2005, when Bartholomeusz et al presented the patterning of different materials using a cutting plotter, and their application in electroplating, shadow masking, PDMS micromolding and laminated microfluidics (Bartholomeusz et al 2005). Since then, a number of microfluidic devices have been implemented using this technique (for example, Nath et al 2014;Pjescic and Crews 2012;Greer et al 2007;Atencia et al 2012;Kido et al 2007;Gorking et al 2010), including several works by our group (Martinez-Duarte et al 2010. The use of xurography to pattern adhesive films has been particularly meaningful to the microfluidics community since robust, inexpensive devices can be fabricated in matter of minutes without the need for a cleanroom.…”

A versatile oscillating‐flow microfluidic PCR system utilizing a thermal gradient for nucleic acid analysis

“…Typically, an additional postprocessing step that takes about 30 min is required by existing NAT devices to obtain melt curves. Previously, we have reported the acquisition of amplification and melt curves using fluorescence images while thermocycling the sample (Crews, Wittwer, & Gale, 2008; Crews, Wittwer, Montgomery, Pryor, & Gale, 2009; Pjescic & Crews, 2012; Pješčić, Tranter, Hindmarsh, & Crews, 2010). This system was based on continuous flow in a thermal gradient, eliminating the need for stringent temperature control.…”

Oscillating-flow Thermal Gradient PCR

“…Typically, an additional post processing step that takes about 30 minutes is required by existing NAT devices to obtain melt curves. Previously, we have reported the acquisition of amplification and melt curves using fluorescence images while thermocycling the sample [15][16][17][18] . This system was based on continuous flow in a thermal gradient, eliminating the need for stringent temperature control.…”