Analytical Ultracentrifuge (AUC)

AUC is the method of choice for determination of the molecular weight of macromolecular complexes (from small polypeptides to viruses) under native solution conditions. The radial distribution of protein in the spinning cell, that arises from the action of both the outward centrifugal force and random diffusion, is mass and shape dependent (in a sedimentation velocity experiment). Analysis of data can determine even very weak association constants and reveal the size distributions of solutions containing multiple species.

We have two Beckman XL-Is have absorption as well as interference optics able to observe sedimenting species under a wide range of buffer conditions and concentrations.

• AUC cell components
• AUC sample preparation
• AUC data analysis software

Differential Scanning Calorimeter (DSC)

Differential Scanning Calorimetry (DSC) measures enthalpy (ΔH) of unfolding due to heat denaturation.  DSC is also used to determine the change in heat capacity. DSC can elucidate the factors that contribute to the folding and stability of native biomolecules, including hydrophobic interactions, hydrogen bonding, conformational entropy, and the physical environment.

The center currently has two Microcal DSCs available for user use.

• DSC sample preparation and instructions
• VP-DSC manual
• VP-DSC origin data analysis tutorial

Isothermal Titration Calorimeter (ITC)

Isothermal Titration Calorimetry (ITC) is a technique that directly measures the heat released or absorbed during a biomolecular binding event. Measurement of this heat enables accurate determination of binding constants, reaction stoichiometry, enthalpy and entropy, thereby providing complete information of the molecular interaction in a single experiment. ITC is the method of choice for characterizing biomolecular interactions.

• ITC instructions
• ITC sample preparation
• ITC review reference

Circular Dichorism (CD)

Circular dichroism (CD) is the property of chiral molecules to absorb the right and left components of circularly polarized light to a different extent. CD spectroscopy measures this differential absorbance.  Nearly all biological macromolecules are chiral and lend themselves to CD experiments. The Aviv Circular Dichroism Spectrometer records CD as a function of wavelength, time and temperature.

The center currently has three Aviv CDs available for user use.

• CD start-up instructions
• CD sample preparation
• CD spectra analysis reference

Fluorescence Spectrometer

Intrinsic fluorescence or site-specific labels can be used to monitor oligomerization, ligand binding, folding and local conformational rearrangements. Fluorescence resonance energy transfer between suitable pairs of fluorophores gives information on their distance. Excitation and emission polarizers allow for anisotropy measurements.

We have two Aviv fluorimeters with titrators available for use.

• Fluorimeter instructions

Fluorescence Plate Reader

We have BMG CLARIOstar plate reader with polarizers that can be used for fluorescence and anisotropy measurements.

Surface Plasmon Resonance

We have a Nicoya open SPR, that can be used to measure binding constants and kinetics.