RESUMO
Quantum dots (QDs) have been used as fluorescent probes in biological and medical fields such as bioimaging, bioanalytical, and immunofluorescence assays. For these applications, it is important to characterize the QD-protein bioconjugates. This chapter provides details on a versatile method to confirm quantum dot-protein conjugation including the required materials and instrumentation in order to perform the step-by-step semiquantitative analysis of the bioconjugation efficiency by using fluorescence plate readings. Although the protocols to confirm the QD-protein attachment shown here were developed for CdTe QDs coated with specific ligands and proteins, the principles are the same for other QDs-protein bioconjugates.
Assuntos
Imunoglobulinas/química , Pontos Quânticos/química , Soroalbumina Bovina/química , Espectrometria de Fluorescência/métodos , Animais , Compostos de Cádmio/química , Bovinos , Humanos , Proteínas Imobilizadas/química , Telúrio/química , Água/químicaRESUMO
The relative stabilities of the alkali [M subset 222]+ cryptates (M = Na, K, Rb and Cs) in the gas phase and in solution (80:20 v/v methanol:water mixture) at 298 K, are computed using a combination of ab initio quantum-chemical calculations (HF/6-31G and MP2/6-31+G*//HF/6-31+G*) and explicit-solvent Monte Carlo free-energy simulations. The results suggest that the relative stabilities of the cryptates in solution are due to a combination of steric effects (compression of large ions within the cryptand cavity), electronic effects (delocalization of the ionic charge onto the cryptand atoms) and solvent effects (dominantly the ionic dessolvation penalty). Thus, the relative stabilities in solution cannot be rationalized solely on the basis of a simple match or mismatch between the ionic radius and the cryptand cavity size as has been suggested previously. For example, although the [K subset 222]+ cryptate is found to be the most stable in solution, in agreement with experimental data, it is the [Na subset 222]+ cryptate that is the most stable in the gas phase. The present results provide further support to the notion that the solvent in which supramolecules are dissolved plays a key role in modulating molecular recognition processes.