Biomedical subjects

A Josten

Publications and source records attributed to A Josten.

3 recordsLinked to original sources

PubMed2005

Oligonucleotides as coding molecules in an anti-counterfeiting system.

Due to the growing numbers of counterfeited products on the world market, there is a huge demand for new and forgery-proof marking systems. We developed a unique system using "molecular beacons" with well adapted thermodynamic parameters. This marking system consists of the three components: DNA tag (a label or directly printing), detection pen (contains the "molecular beacon " solution), and DNA-scanner (reads the specific signal triggered by the detection pen even at daylight). The vast coding capacity of DNA combined with the highly specific signal offers a degree of security that is unmatched by conventional identification technologies.

Biosensing Techniques↗

PubMed2000

Use of microbial transglutaminase for the enzymatic biotinylation of antibodies.

Nowadays many reagents are available for the biotinylation of proteins. As most of them bind to amino groups of the protein the degree of labelling differs from batch to batch and the possibility exists that the biological activity of the target protein may be affected by the labelling procedure. In the present study we have investigated an enzymatic approach to biotinylation using microbial transglutaminase (MTGase) from Streptoverticillium mobaraense. The proposed method is particularly suitable when only a few biotin molecules need to be attached to the target proteins. The enzyme catalyses the acyl transfer reaction between gamma-carboxyamide groups and various primary amines. This was exploited for biotinylation using two amino-modified biotin derivatives, biotinamido-5-pentylamin (BIAPA) and biotinoyl-1,8-diamino-3, 6-dioxaoctane (BIDADOO) as acyl acceptors and a monoclonal IgG against the herbicide 2,4-dichlorophenoxyacetic acid (2,4-D) as the acyl donor. Kinetic studies revealed that the MTGase-mediated reaction proceeds with low velocity and is almost complete after 34 h. Conjugation ratios ranging from 1.1 to 1.9 biotins per IgG were found by mass spectrometry. To investigate the influence of antibody conjugation on antigen binding a competitive ELISA for the determination of 2,4-D employing MTGase-biotinoylated IgGs was developed. In this assay lower limits of detection of 0.3 and 1.0 microg/l of 2,4-D were achieved with BIDADOO- and BIAPA-modified antibodies, respectively.

2,4-Dichlorophenoxyacetic Acid↗

PubMed1998

Microbial transglutaminase-mediated synthesis of hapten-protein conjugates for immunoassays.

Hapten-protein conjugates are essential in many immunochemical assays, in particular, in assays employing titration or competitive assay formats. By exploitation of the catalytic properties of the microbial transglutaminase from Streptoverticillium mobarense sp. (MTGase), i.e., acyl transfer between gamma-carboxamide groups and various primary amines, new techniques for the synthesis of hapten-protein conjugates were developed. This is demonstrated by two examples. The feasibility of MTGase for hapten-protein conjugate synthesis was studied by coupling the herbicide 2,4-dichlorophenoxyacetic acid (2,4-D) to casein. Different procedures for the synthesis and the immobilization of these 2,4-D-casein conjugates were evaluated, comprising (i) a batch procedure, (ii) coupling of 2,4-D to an already immobilized layer of casein, and (iii) a method for simultaneous immobilization and conjugation. Kinetic studies revealed that conjugate formation in the batch procedure was almost complete after approx 2 h. By employing the conjugates in a competitive ELISA, detection limits as low as 0.05 microgram/L 2,4-D were reached. Using the approach with simultaneous immobilization and conjugation, the time for the whole assay could be reduced to only 2 h. Finally, to demonstrate the versatility of the enzymatic synthesis of hapten-protein conjugates, an ELISA for 2,4,6-trinitrotoluene (TNT) determination based on transglutaminase-synthesized conjugates was developed. In this assay, a detection limit as low as 0.04 microgram/l TNT was obtained.

2,4,5-Trichlorophenoxyacetic Acid↗