Fragment capture covalent

Figure 10.1 Principle of covalent capture methods. Drug fragments typically have weak binding affinity and can therefore be difficult to detect. By introducing two reactive groups, X and Y, a fragment that binds in the vicinity of X can be captured covalently by the protein target and easily identified by mass spectrometry.

The array above has 15 rows of spots containing synthetic capture RNA covalently attached to a glass slide. Three spots in each row contain identical capture RNA designed to bind to a short section of one strain of viral RNA. The spot at the left in each row is a control that will become fluorescent in every test and serves as an internal standard. Viral RNA extracted from patients is amplified (reproduced into many copies) and digested (cleaved into fragments). Capture RNA on the slide binds selected viral RNA fragments. Another synthetic RNA with a fluorescent tag is designed to bind to a different section of viral RNA. After allowii d ested viral RNA to bind to capture RNA and to fluorescent RNA, excess fluorescent RNA is washed away. Fluorescence intensity in each spot is related to the amount of viral RNA bound at that spot. 

A two-site immunometric assay of undecapeptide substance P (SP) has been developed. This assay is based on the use of two different antibodies specifically directed against the N- and C-terminal parts of the peptide (95). Affinity-purified polyclonal antibodies raised against the six amino-terminal residues of the molecule were used as capture antibodies. A monoclonal antibody directed against the carboxy terminal part of substance P (SP), covalently coupled to the enzyme acetylcholinesterase, was used as the tracer antibody. The assay is very sensitive, having a detection limit close to 3 pg/mL. The assay is fiiUy specific for SP because cross-reactivity coefficients between 0.01% were observed with other tachykinins, SP derivatives, and SP fragments. The assay can be used to measure the SP content of rat brain extracts. 

As mentioned at the beginning of this review, covalent grafting of biomolecules onto colloidal support avoids reversible immobilization. Furthermore, an extended conformation of the ODNs is a key factor for efficiently hybridizing the complementary DNA fragment (i.e., target capturing). However, implementation of immobilization is more difficult than adsorption. 

T7ery little has appeared in the literature concerning the radiation chemistry of covalent inorganic compounds in condensed phase. In the search for new, high energy oxidizers, it appears plausible that ion fragmentation, electron capture, ion-molecule reactions, and free radical combination reactions at low temperatures may be utilized. 

Figure 1 Template-based synthesis of a macrocycle, showing the formation of the intermediate supramolecular complex between the template and an interacting fragment—in the key equilibrium process—followed by its covalent capture giving the prodnct-tonplate complex, which in this hypothetical case can be dissociated. Note that the template favors the formation of a cyclic tetramo- ovct any other oligomer, cyclic, or linear.

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