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Multiplexed expression and screening for recombinant protein production in mammalian cells

Overview of attention for article published in BMC Biotechnology, December 2006
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About this Attention Score

  • In the top 25% of all research outputs scored by Altmetric
  • High Attention Score compared to outputs of the same age (80th percentile)
  • Average Attention Score compared to outputs of the same age and source

Mentioned by

patent
8 patents

Citations

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39 Dimensions

Readers on

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121 Mendeley
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1 CiteULike
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Title
Multiplexed expression and screening for recombinant protein production in mammalian cells
Published in
BMC Biotechnology, December 2006
DOI 10.1186/1472-6750-6-49
Pubmed ID
Authors

Susan DJ Chapple, Anna M Crofts, S Paul Shadbolt, John McCafferty, Michael R Dyson

Abstract

A variety of approaches to understanding protein structure and function require production of recombinant protein. Mammalian based expression systems have advantages over bacterial systems for certain classes of protein but can be slower and more laborious. Thus the availability of a simple system for production and rapid screening of constructs or conditions for mammalian expression would be of great benefit. To this end we have coupled an efficient recombinant protein production system based on transient transfection in HEK-293 EBNA1 (HEK-293E) suspension cells with a dot blot method allowing pre-screening of proteins expressed in cells in a high throughput manner. A nested PCR approach was used to clone 21 extracellular domains of mouse receptors as CD4 fusions within a mammalian GATEWAY expression vector system. Following transient transfection, HEK-293E cells grown in 2 ml cultures in 24-deep well blocks showed similar growth kinetics, viability and recombinant protein expression profiles, to those grown in 50 ml shake flask cultures as judged by western blotting. Following optimisation, fluorescent dot blot analysis of transfection supernatants was shown to be a rapid method for analysing protein expression yielding similar results as western blot analysis. Addition of urea enhanced the binding of glycoproteins to a nitrocellulose membrane. A good correlation was observed between the results of a plate based small scale transient transfection dot blot pre-screen and successful purification of proteins expressed at the 50 ml scale. The combination of small scale multi-well plate culture and dot blotting described here will allow the multiplex analysis of different mammalian expression experiments enabling a faster identification of high yield expression constructs or conditions prior to large scale protein production. The methods for parallel GATEWAY cloning and expression of multiple constructs in cell culture will also be useful for applications such as the generation of receptor protein microarrays.

Mendeley readers

Mendeley readers

The data shown below were compiled from readership statistics for 121 Mendeley readers of this research output. Click here to see the associated Mendeley record.

Geographical breakdown

Country Count As %
Switzerland 2 2%
United States 2 2%
Italy 1 <1%
Sweden 1 <1%
Brazil 1 <1%
United Kingdom 1 <1%
South Africa 1 <1%
Unknown 112 93%

Demographic breakdown

Readers by professional status Count As %
Researcher 44 36%
Student > Ph. D. Student 22 18%
Student > Master 15 12%
Student > Bachelor 8 7%
Other 8 7%
Other 15 12%
Unknown 9 7%
Readers by discipline Count As %
Agricultural and Biological Sciences 63 52%
Biochemistry, Genetics and Molecular Biology 30 25%
Engineering 5 4%
Chemistry 3 2%
Medicine and Dentistry 3 2%
Other 7 6%
Unknown 10 8%
Attention Score in Context

Attention Score in Context

This research output has an Altmetric Attention Score of 6. This is our high-level measure of the quality and quantity of online attention that it has received. This Attention Score, as well as the ranking and number of research outputs shown below, was calculated when the research output was last mentioned on 02 March 2023.
All research outputs
#4,875,285
of 23,462,326 outputs
Outputs from BMC Biotechnology
#263
of 945 outputs
Outputs of similar age
#21,513
of 158,994 outputs
Outputs of similar age from BMC Biotechnology
#7
of 18 outputs
Altmetric has tracked 23,462,326 research outputs across all sources so far. Compared to these this one has done well and is in the 76th percentile: it's in the top 25% of all research outputs ever tracked by Altmetric.
So far Altmetric has tracked 945 research outputs from this source. They typically receive more attention than average, with a mean Attention Score of 7.8. This one has gotten more attention than average, scoring higher than 65% of its peers.
Older research outputs will score higher simply because they've had more time to accumulate mentions. To account for age we can compare this Altmetric Attention Score to the 158,994 tracked outputs that were published within six weeks on either side of this one in any source. This one has done well, scoring higher than 80% of its contemporaries.
We're also able to compare this research output to 18 others from the same source and published within six weeks on either side of this one. This one is in the 33rd percentile – i.e., 33% of its contemporaries scored the same or lower than it.