Table 2.

Design options for microfluidic implementation of nucleic acid amplification test unit operations.

Unit Operation	Options	POC Issues
Blood Fractionation	centrifugation filtration sedimentation fluid-flow fractionation	Centrifugation is difficult to integrate into the chip, and a separate centrifugation step may be needed. Most LOC filtration/sedimentation and fluid flow fractionation is typically limited to small sample volumes.
Lysis	mechanical chemical enzymatic thermal	Viruses are comparatively easy to lyse with detergents and chaotropic salts; vegetative bacterial cells are somewhat more difficult; and spores most difficult, requiring enzymes and/or mechanical disruption.
Nucleic Acid (NA) Isolation (Extraction, Purification, Concentration)	solid-phase extraction liquid-liquid extraction ethanol precipitation hybridization (e.g., magnetic beads) centrifugation	Solid-phase extraction using a NA-binding phase (e.g., silica) and binding, wash, and elution buffers is readily implemented with microfluidics.
Nucleic Acid (NA) Amplification	PCR (polymerase chain reaction) isothermal methods (e.g., AMP, RPA, HDA, NASBA)	PCR is well developed, but requires instrumentation for precise thermal cycling, and has relatively high power consumption. Isothermal methods require much less instrumentation. LAMP (65 °C constant temperature incubation) appears to be the most used method.
Amplicon Detection	fluorescent dyes bioluminescent reporters colorimetric dyes electrochemical sensors	Fluorescent dyes are very sensitive. Bioluminescent reporters do not require light sources or optical filters. Colorimetric dyes can be read by eye for instrument-free operation. Electrochemical sensors are more compact, but generally less sensitive and difficult to interface with a disposable chip.