We do NOT collect User Data during CMI training sessions.
All Training sessions include:
Overview of the instrument and maintenance procedures
Tutorial on control and analysis software
Review of experimental design
User trainings may be individual or small group, depending on scheduling.
For some technologies, we'll run a standard during training to illustrate how the instrument works and/or to practice. We won't collect any data during training on systems for which sample manipulation is...
The choice of which molecule to keep constant and which to vary will depend on the experiment type, the properties of the molecules and the stoichiometry of the interaction.
For ITC, where neither protein is labeled or immobilized, either molecule can be in the cell or syringe. You will need a higher concentration and total amount of the molecule in the syringe, so solubility and availability will generally be deciding factors. For protein/small molecule interactions, it is most common to put the protein in the cell and the compound in the syringe.
The amount (volume and concentration) of sample that you need will vary by method and by the Kd (equilibrium dissociation constant) of the interaction. For guidelines see the CMI technology comparison table. Binding assays will typically have one binding partner at a fixed concentration (or immobilized on a surface) and one binding partner at a variable concentration. The high concentration for the variable analyte will typically be 10-100X Kd,... Read more about How much sample is needed for a binding assay?
The relative size of the molecules isn't generally the deciding factor. Large proteins will typically give larger signals, but that isn’t necessarily better.
For protein/protein interactions, start by considering other properties of the proteins. Most importantly stoichiometry of binding. If one of the molecules is multivalent (an antibody, for example),...
MST experiments on the Monolith NT.115 pico require that one of the binding partners is fluorescently labeled.
For protein/protein interactions, start by considering the amounts, solubility and sequence of the proteins. The labeled target is typically used at 5-10 nM concentrations (200 ul/titration). The unlabeled ligand is ideally used at a concentration of 100X Kd (equilibrium dissociation constant), so could be uM to mM for weaker binders (20 ul/titration). If one protein is much more soluble and/or more abundant, that may be a better choice for the...
The CMI offers several orthogonal biophysical technologies to measure molecular interactions and to characterize molecular properties. To see how these technologies compare to each other see the Technologies Comparision Tables.
