Novel bioactive molecules can be rationally designed by growing and linking small fragments. Because fragments are fast and promiscuous, it is common to have contradictory hit results between different experimental screening techniques. Here, we simultaneously determine fragment binding poses, affinities, and kinetics on a focused library of 42 fragments against the serine protease factor Xa us...

Three polyaminocyclodextrin materials, obtained by direct reaction between heptakis(6-deoxy-6-iodo)-β-cyclodextrin and the proper linear polyamines, were investigated for their binding properties, in order to assess their potential applications in biological systems, such as vectors for simultaneous drug and gene cellular uptake or alternatively for the protection of macromolecules. In particul...

The aim of binding assays is to measure interactions between two molecules, such as a protein binding another protein, a small molecule, or a nucleic acid. Hard work is required to prepare reagents, but flaws in the design of many binding experiments limit the information obtained. In particular many experiments fail to measure the affinity of the reactants for each other. This essay describes ...

The phenolic pKa of fluorescein varies depending on its environment. The fluorescence of the dye varies likewise. Accordingly, a change in fluorescence can report on the association of a fluorescein conjugate to another molecule. Here, we demonstrate how to optimize this process with chemical synthesis. The fluorescence of fluorescein-labeled model protein, bovine pancreatic ribonuclease (RNase...

Secreted by the adipose tissue, adiponectin is an adipokine with unique anti-diabetic, anti-obesity and antiatherosclerosis properties. Adiponectin has been proposed as a drug target for treating obesity-related diseases and many existing drugs have been found to regulate circulating adiponectin levels in humans. The initial goal of this study was to characterize the binding of potential ligand...

it has been reported that cxcl12 binding to cxcr4 induces several intracellular signaling pathways, and enhances survival, proliferation, and migration of malignant cells. herein we examined the effects of anti-estrogen tamoxifen and anti-allergic tranilast drugs as a single or in combination on invasion by two in vitro invasion assays, wound-healing and matrigel invasion on mcf-7 and mda-mb-23...

ASSAY CURVES, USING A DISK DIFFUSION METHOD FOR THE ANTIBIOTICS GENTAMICIN AND CEFAZOLIN, WERE PREPARED WITH: saline, saline plus 10% serum, and ascitic, synovial, cerebrospinal, and pleural fluids. The curves were compared with a standard curve prepared with pooled human serum. The pH, total protein, glucose, blood urea nitrogen, sodium, potassium, calcium, phosphorus, chloride, CO(2) content,...

the immunogenicity of biotherapeutic molecules, and determine drug concentrations for pharmacokinetic analyses. The accuracy and performance of LBAs, however, depends on the quality of key reagents, called critical reagents. The Global Bioanalysis Consortium (GBC) and the European Medicines Agency classify critical reagents as analyte specific or binding reagents, specifically antibodies; pepti...

In this protocol we demonstrate a method for comparing the competition between GTPase-binding proteins. Such an approach is important for determining the binding capabilities of GTPases for two reasons: The fact that all interactions involve the same face of the GTPases means that binding events must be considered in the context of competitors, and the fact that the bound nucleotide must also b...

The principles of sequential saturation as a form of competitive binding assays are discussed in detail and differentiated from those of equilibrium techniques. The rationale of its applicability in practice and its potential to increase sensitivity under certain assay conditions are demonstrated with three binding systems involving digoxin, insulin, and folates. The advantages and disadvantage...