Performance Evaluation of Fast Microfluidic Thermal Lysis of Bacteria for Diagnostic Sample Preparation

Abstract

Development of new diagnostic platforms that incorporate lab-on-a-chip technologies for portable assays is driving the need for rapid, simple, low cost methods to prepare samples for downstream processing or detection. An important component of the sample preparation process is cell lysis. In this work, a simple microfluidic thermal lysis device is used to quickly release intracellular nucleic acids and proteins without the need for additional reagents or beads used in traditional chemical or mechanical methods (e.g., chaotropic salts or bead beating). On-chip lysis is demonstrated in a multi-turn serpentine microchannel with external temperature control via an attached resistive heater. Lysis was confirmed for Escherichia coli by fluorescent viability assay, release of ATP measured with bioluminescent assay, release of DNA measured by fluorometry and qPCR, as well as bacterial culture. Results comparable to standard lysis techniques were achievable at temperatures greater than 65 °C and heating durations between 1 and 60 s.

Paper-based molecular diagnostic for Chlamydia trachomatis

“…Thermally induced lysis of Gram negative cells is well-established in the use of colony PCR and has been investigated previously in E. coli . 23 These studies by Packard et al . show that greater than 90% of cell viability is lost when E. coli were heated to 60°C for 15 seconds.…”

Time-resolved, single-cell analysis of induced and programmed cell death via non-invasive propidium iodide and counterstain perfusion

“…Conventional staining protocols using PI concentrations higher than 1 μM intended for sorting414, confirmation of cell lysis16, or cellular analytics17181920 have been described for prokaryotes and eukaryotes. PI staining is generally performed as an endpoint measurement, frequently after cell fixation171921.…”

A Microfluidic Platform for Precision Small-volume Sample Processing and Its Use to Size Separate Biological Particles with an Acoustic Microdevice

“…This interface scheme makes for easy chip replacement and rapid device redesign, requiring few or no changes to other system components, provided chip footprints conform to the grid format. For example, we have used this platform with microfluidic chips for continuousflow electrophoresis, thermal cell lysis, 29 rapid mixing of reagents for chemical synthesis, and single-cell capture and interrogation.…”