Tagged antibody

Fluorescent labels, by contrast, can provide tremendous sensitivity due to their property of discrete emission of light upon excitation. Proteins, nucleic acids, and other molecules can be labeled with fluorescent probes to provide highly receptive reagents for numerous in vitro assay procedures. For instance, fluorescently tagged antibodies can be used to probe cells and tissues for the presence of particular antigens, and then detected through the use of fluorescence microscopy techniques. Since each probe has its own fluorescence emission character, more... 

A spectrophotometric assessment of the F/P ratio should be done after purification of the tagged antibody. The measurement of absorbance at 495 nm (for fluorescein) divided by the absorbance at 280 nm should be between 0.3 and 1.0 to obtain a good fluorescent derivative of acceptable activity and low background. This usually translates into a ratio of about 4-7 fluorescein molecules per protein molecule. 

Y. Hirata, M.-L. Laukkanen, K. Keinanen, H. Shigematsu, M. Aizawa, and F. Mizutani, Microscopic characterization of Langmuir-Blodgett films incorporating biosynthetically lipid-tagged antibody. Sens. Actuators B 76, 181-186 (2001). 

E10 MAb to detect myc-tagged antibody fragments (9E10 cell line is available from European Collections of Animal Cell Cultures)... 

For detection of myc-tagged antibody fragments, use 9E10 MAb and HRP-conju-gated monoclonal antimouse IgG Fc secondary antibody. 

Add enzyme-labelled antibody, specific for the target antigen, to the solid phase. The "tagged" antibody will attach to any antigen captured by the solid phase antibody. 

Secondary, magnetic tagging antibody mouse anti PE-MACS microbeads antibody conjugate, isotype IgGl (Miltenyi Biotec, Auburn, CA). 

Fig. 9. Determination of the relative amount of ligand bound per unit protein. The signal intensities from the DNA binding assay (Fig. 6B) have been divided by the relevant anti-His tag antibody binding signal (Fig. 5) to show the relative amount of DNA bound per unit protein for each p53 variant in the array.

Fig. 11. Binding of calmodulin to an array of a diverse set of 144 human proteins enables novel, calcium-dependent interactions to be identified. (A) Binding of Cy3-labeled anti-His tag antibody to the array allows the relative amount of protein in each spot to be determined. (B) Binding of Cy3-labeled calmodulin allows potential interacting partners to be identified. (C) Histogram showing the relative amount of calmodulin bound per unit protein in the presence of calcium ions. (D) Histogram showing the relative amount of calmodulin bound per unit protein in the presence of calcium ions and a high concentration of a divalent metal ion chelator, ethylenediamine tetra-acetic acid.

The method is based on the covalent immobilization of enzyme onto the immunosensor surface, circumventing the use of enzyme-tagged antibody and alleviating the signal instability from low-affinity binding. The electrochemical signal can be maintained even if... 

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