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Storing Mix&Go

How should I store Mix&Go Reagent?
Mix&Go Reagent should be stored at room temperature.
How should I store Mix&Go-activated beads?
We strongly recommend that Mix&Go-activated beads be stored in its own Mix&Go reagent until they are needed for protein coupling. Failure to observe recommendation is likely to compromise the protein-binding capacity of the Mix&Go surface.
How should I store Antibody/Protein-coated Mix&Go beads?
Anteo has developed various storage and coating buffers that have been optimised for each specific bead and protein/antibody. Always follow Anteo’s protocols when coupling and storing your beads. Failure to observe recommendation is likely to compromise the protein-binding capacity of the Mix&Go surface.

Stability of Mix&Go

What is the stability of Mix&Go Reagent?
Mix&Go Reagent can be stored up to 2 years on laboratory bench at room temperature. Please ensure that reagent is sealed properly to avoid contamination.
What is the stability of Mix&Go activated beads?
If you choose not to use Mix&Go activated beads immediately and store for later use, you should:
  • Ensure that the beads do not become contaminated
  • Ensure that they are well sealed
  • Mix&Go activated beads are stable at 4C for up to 2 years
  • Ensure the beads are well dispersed before use
What is the stability of Protein/Antibody Mix&Go activated beads?
If you choose not to use protein/antibody Mix&Go activated beads immediately and store for later use, you should:
  • Ensure that the beads do not become contaminated
  • Ensure that they are well sealed
  • Protein coupled Mix&Go beads are stable at 4C for up to 1 year
  • Ensure the beads are well dispersed before use

Optimizing Mix&Go

How do I know whether I have correctly applied Mix&Go?
Protein loading assay after coupling or specific assay performance will be different from traditional method.
What is the optimal pH and temperature ranges when using Mix&Go?
The pH of Mix&Go solution is 2.8 - 4.2 and Mix&Go-activated beads are stable in this environment. Our data indicates that protein-coupled Mix&Go beads are compatible with commonly used buffers/solutions (with the notable exception of PBS for storage) including:
  • 100 mM MES pH 5.2 and pH 6.0
  • 100 mM MES pH 5.2 and pH 6.0
  • 100 mM Tris pH 8
  • 50 mM acetate-buffered saline pH 5
  • 150 mM normal saline
  • Mix&Go interaction withstands 2% SDS at 100oC for 10 min PBS*
Are there conditions that could be potentially detrimental to a Mix&Go surface or could interfere with its ability to bind?
Yes. During Mix&Go activation there are conditions/agents that are detrimental to the Mix&Go surface (before protein coupling) and can interfere with its ability to bind proteins. These include:
  • Metal chelators such as EDTA
  • Phosphates, including a phosphate-buffered saline (PBS) when using for long term storage NOTE - PBS is not a problem for Mix&Go beads during assay conditions such as assay buffer or washing buffer
  • Charged surfactants

Antibodies and Mix&Go

What concentration of antibody should I use on Mix&Go activated beads?
When coupling antibody to Mix&Go it is recommended that customers perform a titration of their antibody in order to achieve optimal coupling conditions. In the majority of cases when using Mix&Go, the amount of antibody required to perform a given assay can be dramatically reduced. In standard microtiter plates and bead assays, it may be possible to reduce the amount of capture antibody needed for a sandwich immunoassay by as much as 80% and still achieve optimal signal.
Do some antibodies bind better than others to Mix&Go?
Different antibodies have different properties and most would need to be tested empirically and titrated to determine optimal coupling conditions. However, Mix&Go works with the vast majority of antibodies (either polyclonal or monoclonal), even those that can’t be attached using standard chemistries.
Does Mix&Go target Fc regions or will the orientation be somewhat random?
Mix&Go targets a Histadine rich region on the Fc of antibodies and therefore the majority of antibodies are bound in the correct orientation, which results in higher signal and less non-specific binding. We have also shown that due to the generic nature of Mix&Go, antibody fragments (such as Fab and F(ab)2 fragments), Streptavidin, Protein A and G can also bind to the Mix&Go surface with good performance characteristics.
Do Antibody/Protein-coated Mix&Go beads tolerate PBS?
Protein-coated Mix&Go beads do not tolerate storage in PBS for long periods, i.e., more than 2-3 days. As a general rule, protein-coated Mix&Go beads should not be stored in PBS. Always follow Anteo’s protocols when coupling and storing your beads.

Note: PBS is not a problem for Mix&Go beads during assay conditions such as assay buffer.

How Mix&Go Works

What can Mix&Go bind to?
Mix&Go can bind to any group with an accessible electron pair, so groups such as hydroxyl, acid, amine, thiol or phenyl can contribute to the binding. Each binding interaction is relatively weak but because there are multiple binding points, in combination, it forms a very strong interaction.
Does Mix&Go bind to other entities such as antigens?
Mix&Go is able to bind Streptavidin, Protein A and G, and other protein antigens, including small peptides down to 5 kDa, but each protein has to be tested empirically.

Mix&Go Assay Development

Do Mix&Go beads perform well in multiplex assay formats?
We have shown that Mix&Go beads that are coupled with antibody perform well in multiplex immunoassays with no cross talk between individual assays.
What type of assays can be performed with Mix&Go beads?
Sandwich Immunoassay, Antigen-down Immunoassay, Competitive Immunoassay abd Immunoprecipitation assays have all been performed using a Mix&Go bead. There are many more potential applications. The ones listed above are the ones we have most experience with.
Is Mix&Go flexible when it comes to scalability?
Surface coating and protein attachment with Mix&Go can be easily scaled from the milligram to the gram scale.
Is batch-to-batch reproducibility consistent when using Mix&Go coupled beads?
Mix&Go is a polymeric coating and as such can mask inhomogeneity of the underlying surface to a high degree, which greatly improves batch uniformity. Our work has shown that the %CV achieved by three operators simultaneously coupling 8 different antibodies gave an average %CV of 3% for Mix&Go vs. 10% for amide coupling in a sandwich assay.

Mix&Go with Particles/Beads

Do I need any additional reagents when treating beads with Mix&Go?
No. Mix&Go is a “ready to use” reagent. There is no need to defrost or dilute anything prior to your experiment. No need to prepare yourself for a 37 hour protocol! Mix&Go requires only 5min of your time! No need to buy any additional reagents. No need to have any special handling! Just keep the Mix&Go reagent on you lab bench. Mix&Go makes your life easy! The name says it all you just Mix and you Go!
Do magnetic particles have to have carboxyl functionality? Or, could the particles have a plain polystyrene surface?
We are able to activate particles with and without carboxyl groups.
What is the smallest particle diameter that Mix&Go was activated on?
40nm
What is the largest particle diameter that Mix&Go was activated on?
2mm
What beads are compatible with Mix&Go?
Below are just some of the beads that the Mix&Go process works very well with:
  • Ademtech
  • Bangs
  • Bioclone
  • Chemagen
  • Chemicell
  • Cyto-Plex
  • Dynal
  • Dynex
  • JSR
  • Kisker
  • Magspheres
  • Merck/ Estapor
  • Millipore
  • Seradyn
  • Spherotech
  • Varian Lodestar
What beads are not compatible with Mix&Go?
The majority of Anteo’s work has been in the immunoassay space, where we have looked at the performance of Mix&Go technology in limit-of-detection and dynamic-range studies on (typically) acid-modified, polystyrene, magnetic, beads up to 2mm in diameter. We have examined dozens of beads/microspheres/microparticles from a range of companies and we have yet to find a product that is not compatible with Mix&Go!
What is the effect on the beads from various body fluids e.g. blood, serum, and urine?
No effect detected from these fluids on the bead themselves, however the mixtures of proteins in these samples may produce non-specific binding problems, unless the beads are correctly blocked.

Mix&Go Safety Documents

Is there an MSDS of Mix&Go?
Yes. Please email customer.service@oneworldlab.com if you would like to receive it.

Mix&Go Buffer Compatibility

Can Mix&Go coupled beads withstand harsh conditions, such as used in protein purification or immunoprecipitation?
Yes. Mix&Go coupled beads withstand all of the below conditions:
  • Boiling beads for 10 minutes in 2% SDS-PAGE sample buffer (SDS, DTT, Glycerol, Coomassie Blue dye)
  • Treating beads for 48 hours with 0.5M EDTA then boiling them in SDS-PAGE buffer
  • Storing Mix&Go-activated beads in 200mM EDTA for 1 year
  • Extremes of pH ~ 2.5 (Glycine or Citrate buffer elution) to pH ~ 12 (0.5M NaOH washing)
What is the tolerance of Mix&Go coated beads to various organic solvents and chemicals?
Yes. Mix&Go coupled beads withstand all of the below conditions:
  • Boiling beads for 10 minutes in 2% SDS-PAGE sample buffer (SDS, DTT, Glycerol, Coomassie Blue dye)
  • Treating beads for 48 hours with 0.5M EDTA then boiling them in SDS-PAGE buffer
  • Storing Mix&Go-activated beads in 200mM EDTA for 1 year
  • Extremes of pH ~ 2.5 (Glycine or Citrate buffer elution) to pH ~ 12 (0.5M NaOH washing)
What are the effects of Mix&Go beads to high salt or chaotropes?
This is usually not recommended during the coupling process. However following the protein coupling, Mix&Go beads are stable in up to 150 mM salt. Protein-coupled Mix&Go beads tolerate short-term exposure to chaotropic agents up to 300 mM urea.
When eluting protein, does Mix&Go interaction withstand 2% SDS at 100C for 10 min?
Short-term exposure to low pH (2.8) and high pH (10) elution buffers has little or no effect on monoclonal antibody-coated Mix&Go beads. We have done immunoprecipitations with antibody-coupled Mix&Go beads, and looked at both denaturing elutions such as 2% SDS loading buffer at 100C for 10min and non-denaturing elution conditions such as 0.1M Citrate pH 2.5 for 10min. We have observed very pure mouse IgG being eluted and no antibody coming off Mix&Go beads when low pH elution is used.

Blocking with Mix&Go

Do I need to block?
In most cases unblocked Mix&Go coated beads have very low non-specific binding. However, we recommend blocking in order to reduce non-specific binding that can lead to high backgrounds. Non specific binding of other proteins to unoccupied spaces on the surface can be disadvantageous to the overall specificity and therefore sensitivity of your assay results.

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Further Technical Assistance
For additional technical support or product information, please contact One World Lab at customer.service@oneworldlab.com or (855) OWL-LABS.