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Protein Stability Analysis

Fluorescence Spectrometry

Our Analytical services include Fluorescence spectroscopy, which is an important investigational tool in many areas of analytical science, due to its extremely high sensitivity and selectivity. We offer services with a Perkin Elmer LS 55 fluorescent spectrophotometer with a thermo regulated cell holder. We provide static fluorescence analysis - intrinsic as well extrinsic fluorescence at controlled thermal environment.

Fluorescence techniques are offered to obtain thermodynamic and kinetic information about transitions of macromolecules, such as protein folding reactions. In studies with proteins, the fluorophores can be either intrinsic (tryptophan, tyrosine residues, co-enzymes) or extrinsic (attached dansyl, fluorescein, pyrene) probes.

Near & Far UV Circular dichroism (CD) and CD with thermal denaturation

Circular dichroism (CD) spectroscopy measures differences in the absorption of left-handed polarized light versus right-handed polarized light which arise due to structural asymmetry, and is indicative of protein structure. The secondary structure can be determined by CD spectroscopy in the "far-UV" spectral region and that in the “near-UV" spectral region can be sensitive to certain aspects of tertiary structure.

Very many times, it is necessary to demonstrate that different lots of a protein have equivalent conformations, for example after a scale-up in the purification process or to qualify a new manufacturing site, and CD can be a good tool for this. Also, such tools are very useful for comparability studies between different products.

Thermal stability of proteins is also assessed using CD by following changes in the spectrum with increasing temperature.

Differential Scanning Calorimetry

Differential Scanning Calorimetry (DSC) is a powerful analytical tool which directly measures the stability and unfolding of a protein. In DSC, the biomolecule is heated at a constant rate and there is a detectable heat change associated with thermal denaturation. DSC measures ΔH of unfolding due to heat denaturation.  The transition midpoint Tm is the temperature where 50% of the protein is in its native conformation and the other 50% is denatured. The higher the Tm, the more stable the molecule.  Such tools are very useful in stability studies and formulation studies as well as product comparability studies.