Label-free assays

Fig. II.8 Assays for 10 ng/mL PSA on IL Class BioCDs. (a) Forward label free assay at 10 ng/mL showing the histrogram of approximately 10,000 spots pairs, (b) Sandwich assay shows clear separation...

L. Label-free assays on the bind system. J. Biomol. Screen. 2004, 9, 481-490. 

Fig. 5.15 Analytical set-up for on-line label-free assay based on ESI-MS. MS instrument Ion-trap mass spectrometer (LCQ Deca, Thermo Electron). PI Carrier/HPLC pump. P2 HPLC pump delivering receptor solution. P3 HPLC pump delivering dissociation solution. PA HPLC pump for final LC-MS analysis of released ligands. 1 Mixing union. 2 Microcoil reactor. VI injection valve.

Cunningham BT, Li P, Schulz S, Lin B, Baird C, Gerstenmaier J, Genick C, Wang F, Fine E, Laing L (2004) Label-free assays on the BIND system. J Biomol Screen 9 481-490... 

Kelvin nanoprobe (63) are some of the tools that have been adopted for label-free assay systems. 

DNA oxidation at carbon electrodes is associated with the irreversible oxidation of guanine and adenine [33], For example, the G oxidation signal observed at -i-1.0 V, without external labels, has been used to monitor telomerase activity by using a carbon graphite electrode (CGE) as an electrochemical transducer [5], Telomerase activity has been detected in cell extracts containing as low as 100 ng pl of protein. This label-free assay is practical in the quantitative determination of telomerase activity providing a cheap and simple detection protocol for the diagnosis of cancer that can also be extended to the analysis of food related to DNA. 

Common DNA recognition protocols rely on thiolated ONs adhered to gold surfaces as recognition ligands. Following PCR amplification of DNA to increase assay sensitivity, samples are exposed to the biointerface with the immobilized ON. While label-free assays are possible [125], the majority... 

Rothmund M., Schtitz A., Brecht A., Gauglitz G., Berthel G., Graefe D., Label free binding assay with spectroscopic detection for pharmaceutical screening, Fresenius J Anal Chem 1997 359 15-22. 

Birkert O., Gauglitz G., Development of an assay for label-free high-throughput screening of thrombin inhibitors by use of reflectometric interference spectroscopy, AnalBioanal Chem 2002 372 141-147. 

On the basis of Scheme 5, we recently realized label-free sequence-specific DNA detection with SNP selectivity with the aid of SI nuclease [59]. In this assay, IBr and TO are chosen as the energy donor and acceptor, respectively. 

Fig. 5.14 Principle of label-free ligand binding MS assays. Protein (P) molecules react with the test ligand (L) to form a protein-ligand complex (PL). Unbound compounds are separated from PL by passage through a restricted-access column. Subsequently, PL is dissociated at low pH, and active ligands L are detected by LC-ESI-MS.

Flow Injection Label-free MS Assay Screening of Natural Extracts... 

Fig. 5.17 Demonstration of MS-based bioassay functionality using a plant extract. MS instrument Ion-trap mass spectrometer (LCQ Deca, Thermo Electron), (a) MS analysis of pure extract by direct injection onto restricted-access column 2 in the absence of affinity protein, (b) Analysis of the same natural extract spiked with digoxin using the label-free MS assay method as shown in Fig. 5.15.

A label-free comprehensive platform for functional evaluation of endogenous receptors. Assay and Drug Development Technologies, 4 (5), 609-619. 

Surface plasmon resonance (SPR) technique had become popular in interaction studies between biological molecules (1). It is an optical biosensor, and the interactions can be detected by SPR angle shift or reflection light intensity. In typical SPR measurement, one of pair interacting biomolecules was immobilized on a gold chip, and another was flowed over the chip as its solution. There are two major advantages in SPR assay (a) real time evaluations on kinetics studies and (b) label-free measurements. 

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