On-line cell lysis of bacteria and its spores using a microfluidic biochip

Cíchová, Marianna; Prokšová, Miloslava; Tóthová, Lívia; Sántha, H.; Mayer, Viktor

doi:10.2478/s11535-012-0005-8

Cited by 5 publications

(3 citation statements)

References 30 publications

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“…Lysozyme digests the peptidoglycan by breaking the glycosidic bond, while EDTA protects against DNA degradation by inhibiting the activity of cellular enzymes [ 33 ] by binding with metallic cations. Cichova et al [ 34 ] developed a continuous flow biochip for online bacterial cell lysis and DNA extraction using 1% Triton X-100 and 5% Chelex. Chelex was added to the lysis buffer to protect the DNA from degradation by the DNases.…”

Section: Lysis Techniquesmentioning

confidence: 99%

Towards Multiplex Molecular Diagnosis—A Review of Microfluidic Genomics Technologies

Basha

Yousuff

et al. 2017

Micromachines

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Highly sensitive and specific pathogen diagnosis is essential for correct and timely treatment of infectious diseases, especially virulent strains, in people. Point-of-care pathogen diagnosis can be a tremendous help in managing disease outbreaks as well as in routine healthcare settings. Infectious pathogens can be identified with high specificity using molecular methods. A plethora of microfluidic innovations in recent years have now made it increasingly feasible to develop portable, robust, accurate, and sensitive genomic diagnostic devices for deployment at the point of care. However, improving processing time, multiplexed detection, sensitivity and limit of detection, specificity, and ease of deployment in resource-limited settings are ongoing challenges. This review outlines recent techniques in microfluidic genomic diagnosis and devices with a focus on integrating them into a lab on a chip that will lead towards the development of multiplexed point-of-care devices of high sensitivity and specificity.

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Section: Lysis Techniquesmentioning

confidence: 99%

Towards Multiplex Molecular Diagnosis—A Review of Microfluidic Genomics Technologies

Basha

Yousuff

et al. 2017

Micromachines

View full text Add to dashboard Cite

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“…These particle and matrix associated approaches are more suitable for miniaturization than the traditional chloroform-phenol extraction [6]. Various chip gadgets have been developed for preparation of DNA from bacteria [7], [8], [9], [10], [11]. Silica beads [7], [8] with superparamagnetic iron oxide cores [9] are also very common for on-chip application.…”

Section: Introductionmentioning

confidence: 99%

Evaluation of a microfluidic chip system for preparation of bacterial DNA from swabs, air, and surface water samples

et al. 2016

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The detection of bacterial pathogens from complex sample matrices by PCR requires efficient DNA extraction. In this study, a protocol for extraction and purification of DNA from swabs, air, and water samples using a microfluidic chip system was established. The optimized protocol includes a combination of thermal, chemical and enzymatic lysis followed by chip-based DNA purification using magnetic particles. The procedure was tested using Gram-positive Bacillus thuringiensis Berliner var. kurstaki as a model organism for Bacillus anthracis and the attenuated live vaccine strain of Francisella tularensis subsp. holarctica as Gram-negative bacterium. The detection limits corresponded to 103 genome equivalents per milliliter (GE/ml) for surface water samples spiked with F. tularensis and 102 GE/ml for B. thuringiensis. In air, 10 GE of F. tularensis per 10 L and 1 GE of B. thuringiensis per 10 L were detectable. For swab samples obtained from artificially contaminated surfaces the detection limits were 4 × 103 GE/cm2 for F. tularensis and 4 × 102 GE/cm2 for B. thuringiensis. Suitability of the chip-assisted procedure for DNA preparation of real samples was demonstrated using livestock samples. The presence of thermophilic Campylobacter spp. DNA could be confirmed in air samples collected on pig and broiler farms.

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“…6 Buffers such as ammonium chloride, known as Red Blood Cell Buffer, 7 detergents (disrupting the cell membranes by solubilizing the membrane proteins and lipids, thus creating pores) including Triton X-100 8 and sodium dodecyl sulfate (SDS), 9 chaotropic agents like guanidinium thiocyanate (GTC), 9 ethanol and magnesium chlo-ride (interfering with intermolecular forces in proteins) and enzymes such as lysozyme, 10,11 lysostaphin and proteinasek11 (digesting the peptidoglycans in the cell walls, ultimately compromising their integrity) are the most common chemical agents used. Possible interference of the chemical and enzymatic agents used with the subsequent analysis processes is the major downside of chemical lysis.…”

Section: Introductionmentioning

confidence: 99%

Lysis of gram-positive and gram-negative bacteria by antibacterial porous polymeric monolith formed in microfluidic biochips for sample preparation

2014

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Bacterial cell lysis is demonstrated using polymeric microfluidic biochips operating via a hybrid mechanical shearing/contact-killing mechanism. These biochips are fabricated from a cross-linked poly(methyl methacrylate) (X-PMMA) substrate by well-controlled, high-throughput laser micromachining. The unreacted double bonds at the surface of X-PMMA provide covalent bonding for the monolith, thus contributing to the mechanical stability of the biochip and eliminating the need for surface treatment. The lysis efficiency of these biochips was tested for gram-positive (Enterococcus saccharolyticus and Bacillus subtilis) and gram-negative bacteria (Escherichia coli and Pseudomonas fluorescens), and confirmed by off-chip PCR without further purification. The influence of the flow-rate when pumping the bacterial suspension through the monolith, and of the hydrophobic-hydrophilic balance on the cell lysis efficiency was investigated at a cell concentration of 10 5 CFU/ml. The lysis efficiency of the biochips was better than for off-chip chemical, mechanical, and thermal lysis techniques. The biochip also acts as a filter that isolates cell debris and allows PCR-amplifiable DNA to pass through. As expected, the system performs more efficient lysis for gram-negative than for gram-positive bacteria. The biochip does not require chemical/enzymatic reagents, power consumption, or complicated design and fabrication processes, which makes it an attractive on-chip lysis device that can be used in sample preparation for bio-genetics and point-of-care diagnostics. The biochips were reused for 20 lysis cycles without significant performance degradation or carryover when they were back-flushed between cycles. The biochips efficiently lysed both gram-positive and gram-negative bacteria in about 35 min per lysis and monolith regeneration cycle.

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On-line cell lysis of bacteria and its spores using a microfluidic biochip

Cited by 5 publications

References 30 publications

Towards Multiplex Molecular Diagnosis—A Review of Microfluidic Genomics Technologies

Towards Multiplex Molecular Diagnosis—A Review of Microfluidic Genomics Technologies

Evaluation of a microfluidic chip system for preparation of bacterial DNA from swabs, air, and surface water samples

Lysis of gram-positive and gram-negative bacteria by antibacterial porous polymeric monolith formed in microfluidic biochips for sample preparation

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