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Experimental Strategies
Published in Clive R. Bagshaw, Biomolecular Kinetics, 2017
In the absence of any useful intrinsic absorption or fluorescence signal from the protein, several other strategies can be used to monitor interactions optically. The protein can be labeled with an extrinsic chromophore (or fluorophore) or the spectral properties of the ligand (or an analog) can be used. Alternatively an indicator dye may be used. Many reactions are accompanied by a pH change, which can be exploited to follow the reaction. In addition, protein–protein interactions that involve hydrophobic regions can often be monitored by displacement of a weak-binding aromatic fluorophore, such as ANS (8-anilino-1-naphthalenesulfonate) [344,345]. Hydrophobic probes are also widely used to follow protein folding reactions in which intermediates display accessible hydrophobic regions [346].
Self-assembled non-covalent protein-drug nanoparticles: an emerging delivery platform for anti-cancer drugs
Published in Expert Opinion on Drug Delivery, 2020
Islam A. Hassanin, Ahmed O. Elzoghby
Silk and elastin-like protein polymers (SELP) are recombinant, synthesized protein polymers which offer the capacity for self-assembly, for the delivery of hydrophobic drugs. It was found that the self-assembly of these materials was mediated by electrostatic interactions which were concentration-dependent [74]. SELP NPs were designed to encapsulate hydrophobic drugs. 1-anilino-8-naphthalenesulfonate (ANS), a hydrophobic molecule, was found to induce the formation of micelle-like structures and hence the encapsulation of ANS, which was driven by the hydrophobic interactions between the protein and ANS. Additionally, it was found that doxorubicin (DOX) induced the self-assembly of SELP NPs (Table 6). However, on adding the water-soluble DOX hydrochloride rather than the free DOX base, the loading of DOX was decreased, indicating the role of hydrophobic interactions in the formation of these NPs. Moreover, the recombinant SELPs, which were prepared of silk to elastin block ratios of 1:8, showed the least critical micellar concentration (CMC) at 0.125 mg/mL and the highest loading efficiency of 6.5% for Dox [23].
Assessment of structural and functional similarity of biosimilar products: Rituximab as a case study
Published in mAbs, 2018
Neh Nupur, Nidhi Chhabra, Rozaleen Dash, Anurag S. Rathore
Intrinsic fluorescence spectroscopy was performed on a Spectra Max M2e Multimode Microplate Reader (Molecular Devices) spectrophotometer with a path length of 1 cm. Emission spectra of the samples were recorded at temperature of 25°C in the range of 300–400 nm with an excitation wavelength of 280 nm. The slits for excitation was 5 nm and emission was 10 nm. All samples were diluted to 0.2 mg/mL with sample buffer. Furthermore, extrinsic fluorescence spectroscopy was employed using fluorescent dye, 1-anilino-8-naphthalenesulfonate (ANS), to examine hydrophobic milieu such as protein interiors. Emission spectra of the samples were recorded in the range of 400–600 nm with an excitation wavelength of 280 nm.