Cell Disruption ofEscherichia coliby Glass Bead Stirring for the Recovery of Recombinant Proteins

Song, Donna D.; Jacques, Nicholas A.

doi:10.1006/abio.1997.2149

Cited by 25 publications

(23 citation statements)

References 3 publications

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“…In addition, E. coli is often used for the production of recombinant proteins. The first step in recovering intracellular proteins is cell disruption (Song and Jacques, 1997). Various devices for cell disruption by ultrasonication, shearing, or bead milling are available.…”

Section: Introductionmentioning

confidence: 99%

Disrupting Escherichia coli: A Comparison of Methods

Benov¹,

Al-Ibraheem²

2002

BMB Reports

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The often-encountered problem of disrupting bacteria for the purpose of extracting soluble protein has generated various methods. Many require specialized equipment. Very often, especially during preliminary studies, investigators need a simple, fast, and inexpensive method for cell disruption that preserves biological activity. This paper compares some simple and inexpensive methods for cell disruption, such as bead-vortexing, freezing-thawing, French pressing, and sonication. It also provides some tips to increase protein yield and preserve biological activity. If performed under optimal conditions, bead-vortexing gives protein yields that are comparable to French pressing and sonication. It also preserves the activities of labile enzymes and releases periplasmic enzymes. Vortexing with glass beads appears to be the simplest method for cell disruption.

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Section: Introductionmentioning

confidence: 99%

Disrupting Escherichia coli: A Comparison of Methods

Benov¹,

Al-Ibraheem²

2002

BMB Reports

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“…LTCC unsupported bridges have been used to carry thick film structures for rapid thermal response [17], coils [ 18 ]and for an air core microstrip. [19] Suspended thick films have been fabricated that would enhance both of these applications. [20] Thick film diaphragms have also been demonstrated, [20,21] as have thick conductor heat-spreading columns and tapes in cooling channels.…”

Section: Materials and Propertiesmentioning

confidence: 99%

“…This step is complicated by the wide variability in the susceptibility of biological agents to lysis and solubilization [13]. As a result, a variety of techniques have been developed for the lysis of bacterial agents including chemical and detergent lysis [14], [15], enzyme treatment [16], sonication [17], heating [18], and glass bead milling [19]. Bacterial spores are extremely resistant to lysis and solubilization, and typically require a combination of these aforementioned techniques to prepare them for analysis [20], [21].…”

Section: B High Temperature Lysermentioning

confidence: 99%

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Macro-meso-microsystems integration in LTCC : LDRD report.

Smet¹,

Nordquist²,

Turner³

et al. 2007

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Low Temperature Cofired Ceramic (LTCC) has proven to be an enabling medium for microsystem technologies, because of its desirable electrical, physical, and chemical properties coupled with its capability for rapid prototyping and scalable manufacturing of components. LTCC is viewed as an extension of hybrid microcircuits, and in that function it enables development, testing, and deployment of silicon microsystems. However, its versatility has allowed it to succeed as a microsystem medium in its own right, with applications in non-microelectronic meso-scale devices and in a range of sensor devices. Applications include silicon microfluidic 'chip-and-wire' systems and fluid grid array (FGA)/microfluidic multichip modules using embedded channels in LTCC, and cofired electro-mechanical systems with moving parts. Both the microfluidic and mechanical system applications are enabled by sacrificial volume materials (SVM), which serve to create and maintain cavities and separation gaps during the lamination and cofiring process. SVMs consisting of thermally fugitive or partially inert materials are easily incorporated. Recognizing the premium on devices that are cofired rather than assembled, we report on functional-as-released and functional-as-fired moving parts. Additional applications for cofired transparent windows, some as small as an optical fiber, are also described. The applications described help pave the way for widespread application of LTCC to biomedical, control, analysis, characterization, and radio frequency (RF) functions for macro-meso-microsystems. Tables Executive SummaryUpon the recognition that LTCC technology can play an important part in integration of macro-, meso-, micro-, and nano-scale systems, several areas have been investigated. These include new sacrificial volume materials/methods (SVM), new structures, creative use of thermistors, and setter and jacketed setter materials to fabricate structures never before envisioned. These include moving parts that are self-assembling at the mesoscale. These parts have been actuated electrostatically, pneumatically, and directly mechanically. 6March 2007 In a sense, almost every use of solid SVM is crying out for a patterned definition. Many more ways to do this exist when the SVM is in the form of a fluid. SVMs have no real minimum thickness with respect to the scale of features envisioned for thick films. We have also investigated the upper end of features, successfully fabricating channels and volumes with mm scale dimensions. By using composite SVMs, there is essentially no limit.We have fabricated stacked parts in the form of washers of diameter approximately 1.2 cm. Evaluations have been performed on similar parts that are inches in diameter.The application of sensitive materials has made it possible to fabricate smart channels. The bulk of this work has been carried out with polymeric chemiresistors, but new promise is held for cofirable sensitive materials as well. The two that were examined in detail include thermoresistive and piezor...

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“…After the fermentation step, a sequence of operations is followed to recover pDNA molecules from the host cells and to remove impurities and contaminants from the lysate, until the desired level of purity and other specifications are obtained [4]. Cell disruption is the first and the most critical step in the bioprocess since this step influences the yield and quality of the product [5,6]. The cell disruption method for plasmid isolation must be chosen such that minimal damage is caused on the pDNA product, and in most cases, it is also desired to avoid shearing of the host cell genomic DNA (gDNA) into smaller fragments that are more difficult to separate from pDNA [7].…”

Section: Introductionmentioning

confidence: 99%

Efficient Disruption of Escherichia coli for Plasmid DNA Recovery in a Bead Mill

et al. 2017

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Featured Application: In general, the alkaline lysis is more acceptable than the mechanical lysis for pDNA recovery due to the risk of product degradation when the latter is used. In this work, the experimental design allowed us to explore mechanical disruption conditions under which such an effect was minimized. The application of this technique is important for novel pDNA-vaccine process development, since bead milling is one of the most preferable mechanical cell lysis methods on an industrial scale due to its scalability, ease of operation, controllability, and ability to load concentrated cell slurry. Abstract:The release kinetics of pDNA in a bead mill was studied. Samples taken during the process were analyzed to determine total pDNA (pDNA(t)) and supercoiled pDNA (pDNA(sc)) concentration. In order to identify important variables of the process and to develop an empirical model for optimal pDNA(t) and pDNA(sc) release, a two level 2 3 factorial design was used with variables: mill frequency, cell concentration, and bead size. The results were analyzed by response surface methodology. The optimized conditions for pDNA(t) yield 13.26 mg/g dcw (93.41% recovery), with a mill frequency of 30 Hz, a bead size of 0.10-0.25 mm, and a cell concentration of 20 g wcw/L. However, the optimized conditions for pDNA(sc) yield 7.65 mg/g dcw (92.05% recovery), with a mill frequency of 15 Hz, a bead size of 0.10-0.25 mm, and a cell concentration of 10 g wcw/L. Cell disruption in a bead mill was proved efficient for the release of pDNA(t) and pDNA(sc) compared to the alkaline treatment. The results obtained suggest a compromise between pDNA(sc) purity and recuperation in the process development.

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Cell Disruption ofEscherichia coliby Glass Bead Stirring for the Recovery of Recombinant Proteins

Cited by 25 publications

References 3 publications

Disrupting Escherichia coli: A Comparison of Methods

Disrupting Escherichia coli: A Comparison of Methods

Macro-meso-microsystems integration in LTCC : LDRD report.

Efficient Disruption of Escherichia coli for Plasmid DNA Recovery in a Bead Mill

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