Proten Analysis about

The Protein Analysis Unit provides Biomolecular interaction analysis (BIA), Isothermal Titration Calorimetry (ITC) and Differential Scanning Calorimetry (DSC) , Microscale Thermophoresis (MST)  and NanoSight to scientists at the Weizmann Institute and in Israel in general.

Biomolecular interaction analysis (BIA), Biacore

This biosensor technology is based on the principle of surface plasmon resonance (SPR). This phenomenon occurs when polarized light, under conditions of total internal reflection, strikes an electrically conducting gold layer at the interface between media of different refractive index: the glass of a sensor surface (high refractive index) and a buffer (low refractive index). SPR is a powerful technique for measuring binding between all types of molecular interactions. This includes drugs and targets, antibodies and antigens or any pair of interacting molecules including protein-protein, protein-nucleic acid and protein-lipid interactions. Interactions are measured in real time allowing relative comparisons between different molecules or complete determination of kinetic parameters. SPR does not require any type of labelling of the test molecules Real time kinetics data is accomplished on the our Unit BIAcore T200 instrument. Quantitative information, such as kinetic parameters and equilibrium constants for complex formation can be obtained within a range of 100 m M-1 pM are analyzed with the BIAcore evaluation software. Biacore systems characterize molecules in terms of their:

  • specificity of their interactions
  • on and off rates constants (kinetics)
  • binding strength (affinity)


Isothermal Titration Calorimetry (ITC)

ITC is a powerful analytical tool which measures the binding affinity and thermodynamics between any two biomolecules. Binding constants in the range of 103 to 108 M-1 can be accurately measured.
ITC is considered the “gold standard” assay for binding, and has many advantages:

  • Universal assay – directly measures heat change associated with binding
  • Label-free - uses native materials
  • True in-solution technique
  • Requires minimal assay development
  • Has no molecular weight limitations
  • Non-optical
  • Versatile, can be used with any biomolecule - proteins, nucleic acids, small molecule drugs, lipids, etc.
  • Can be used with a wide range of biological buffers, ionic strengths, pH’s

In a single ITC experiment, one can determine:

  • Binding affinity - Kd
    • Directly measure sub-millimolar to nanomolar
    • Can extend affinity range to picomolar using competitive ITC method
  • Number of binding sites
  • Multiple and different binding sites
  • Enthalpy (ΔH) and entropy (ΔS) of binding

Differential Scanning Calorimetry (DSC)

DSC continuously measures heat capacity of a system as a function of temperature allowing for simultaneous determination of enthalpy, entropy and Gibbs energy for a thermal transition. This information has proven to be extremely important in the characterization of protein folding and oligonucleotide conformation.
Differential Scanning Calorimetry (DSC) is a powerful analytical tool which directly measures the change in heat associated with the unfolding of a protein, lipid, or nucleic acid.  In DSC, as the biomolecule is heated at a constant rate, a detectable heat change associated with thermal denaturation can be accurately measured. DSC experiments are:

  • Label-free and use native materials
  • True in-solution technique
  • Easy to perform - require minimal assay development
  • Conducted using a wide range of biological buffers, ionic strengths and pH’s
  • Universal assay – measures the heat change associated with denaturation
  • Non-optical – unaffected by colored or turbid samples
  • Versatile - can be used with proteins, nucleic acids, lipids and other biomolecules

In a single DSC experiment, one can determine:

  • Transition midpoint (Tm)
  • Enthalpy (ΔH) and heat capacity change (∆Cp) associated with unfolding

Microscale Thermophoresis (MST)

Microscale thermophoresis (MST) is an easy to handle new technology for the analysis of the interaction of biomolecules. MST is a rapid and precise method to quantify biomolecular interaction in solution at microliter scale. Microscale thermophoresis is the directed movement of particles in a microscopic temperature gradient. Any change of the hydration shell of biomolecules due to changes in their structure/conformation results in a relative change of movement along the temperature gradient and is used to determine binding affinities, binding kinetics and activity kinetics. MST allows to measure biomolecule interactions (small molecules, DNA, RNA, proteins, peptides, sugars, lipids, ribosomes etc.) under close-to-native conditions directly in solution without the need of immobilization to a surface (immobilization-free technology). The measurement can be performed in a solution of choice, ranging from standard and proprietary buffers to complex bioliquids including blood serum or cell lysates. One of the reactant should be fluorescently labeled. MST instrument Monolith NT.115 allows to measure the binding of all kinds of biomolecules. The range of reliable measurements suitable for KD values from 10-12 to 10-3 M) as well as the activity of enzymes.


Nanosight instrument NS300 is used for rapid, automated analysis of the size distribution and concentration of all types of nanoparticles from ~10nm to 1000nm in diamter.