If your sample is not challenging you, then your sample might not be challenging (enough)!

 

ProteoPlex MacroDSF instrument 

 

With recent technological advances in X-ray and EM technology, the specimen itself has often become the limiting factor. In EM, high-end transmisson electron microscopes and direct electron detectors are in widespread worldwide use. Astoundingly we do not yet see the same technological push and automation in the one crucial step ahead - sample preparation. Microscope and detectors cannot improve a bad specimen. Many people might say they do not have a problem with their sample and rely on conventional Thermofluor / DSF for sample optimization. We postulate that when there is no problem with the sample, the sample is too easy!

 

ProteoPlex MacroDSF guides you to more stable proteins and thus improves specimen quality - even and especially for complex proteins. ProteoPlex MacroDSF helps you find the ideal conditions - optimal protein concentration, pH value, salt, buffer, additive(s), etc. - to stabilize your protein. This knowledge can dramatically improve your purification results and allows high quality structure determination - e.g. your protein might regularly crystallize after MacroDSF improvement. Due to its unique biophysical algorithm, ProteoPlex MacroDSF is the only technology that can cope with protein complexes that show multi-state unfolding behavior. 

ProteoPlex 001

MPIBPC,

Goettingen,

Germany

ProteoPlex 002

NovAliX SAS,

Illkirch, France

ProteoPlex 003

University of Minnesota,

The Hormel Institute,

Austin, MN, USA

Core parts of ProteoPlex MacroDSF

  • Hamilton liquid handling robot
  • Bio-Rad RT-PCR
  • Thermo well plate sealing machine (optional)
  • Patented ProteoPlex MacroDSF software for multi-state unfolding analysis

 

Functionality

  • Dilution screen: screen for optimal dilution to achieve low protein consumption and best results
  • Buffer screen: screen 88 buffer conditions simultaneously
  • Additive screen: screen 88 additives simultaneously
  • Obtain ranking of best results

Advantages

  • Speed: screen 88 buffer conditions in less than 2 hours
  • Reproducibility: rule out most significant sources of errors (pipetting volume, sample contamination, air bubbles, suboptimal dilution, etc.)
  • Multiple unfolding analysis: only technology that can correctly interpret multi-state unfolding (and of course also two-state unfolding) --> particularly in early purification stages no overlap between ProteoPlex and DSF results!

 

Suggested further reading

Chari, A.; Haselbach, D.; Kirves, J. M.; Ohmer, J.; Paknia, E.; Fischer, N.; Ganichkin, O.; Moller, V.; Frye, J. J.; Petzold, G. et al.: ProteoPlex: Stability optimization of macromolecular complexes by sparse-matrix screening of chemical space. Nature Methods 12 (9), S. 859-865 (2015)
Download: http://www.nature.com/nmeth/journal/v12/n9/full/nmeth.3493.html

 

User manual

ProteoPlex MacroDSF User Manual v06
20180515 ProteoPlex MacroDSF Instruction[...]
PDF-Dokument [6.8 MB]

MacroDSF Flyer

Flyer MacroDSF
20170113 ProteoPlex flyer v01.pdf
PDF-Dokument [621.2 KB]

Important remarks, caveats & limitations

  • ProteoPlex MacroDSF is not applicable to membrane proteins​ as the micelles distort the signal
  • Buffer improvement does not always mean to get better images. There are other parameters that can destabilize or even destroy a protein complex. Thus, buffer optimization is not a panacea but one of many levers to be checked; it can be a limiting condition - or not.
  • Yes, there are cases where Thermofluor/DSF yields similar results as ProteoPlex MacroDSF. But, as a matter of fact, Tm is a physically wrong criterion for complexes which might - coincidentally - be "right". In general, the results from MacroDSF and Thermofluor/DSF converge with increasing protein optimization, i.e. results of both methods agree for complexes that require no further optimization and are already present in a stabilized form. In the early stages of a project, there should hardly be any overlap between the two methods and that is where ProteoPlex can be an extraordinary asset, as shown in Fig.1.

Fig. 1: Comparison to conventional data interpretation. Comparison of melting-curve evaluation by ProteoPlex and conventional DSF. Results obtained by both methods perfectly agree (green) only for complexes that require no further optimization and are already present in a stabilized form. In early stages of a project, there is no overlap between DSF and ProteoPlex results.

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Still want to read more?

Joint press release with NovAliX, 27th March 2018

"NovAliX enters into cooperation agreement with ProteoPlex"

Press release EMBL,

4th Aug 2016:

"Every atom counts"

EMBL Science,

8th Aug 2016:

"Blocking the cell's waste disposal unit"