Biotin-streptavidin

In an extensive SFA study of protein receptor-ligand interactions, Leckband and co-workers [114] showed the importance of electrostatic, dispersion, steric, and hydrophobic forces in mediating the strong streptavidin-biotin interaction. Israelachvili and co-workers [66, 115] have measured the Hamaker constant for the dispersion interaction between phospholipid bilayers and find A = 7.5 1.5 X 10 erg in water. 

Lee G U, Kidwell D A and Colton R J 1994 Sensing discrete streptavidin-biotin interactions with atomic force microscopy Langmuir 10 354... 

Grubmiiller et al., 1996] Grubmiiller, H., Heymann, B., and Tavan, P. Ligand binding and molecular mechanics calculation of the streptavidin-biotin rupture force. Science. 271 (1996) 997-999... 

Both the AFM rupture experiments as well as our simulation studies focussed on the streptavidin-biotin complex as a model system for specific ligand binding. Streptavidin is a particularly well-studied protein and binds its ligand biotin with high affinity and specificity [51]. Whereas previous experiments (see references in Ref. [49]) and simulation studies [52] referred only to bound/unbound states and the associated kinetics, the recent AFM... 

In summary, our simulations provided detailed insight into the complex mcf hanisms of streptavidin-biotin rupture. They attribute the binding force... 

Fig. 6. Force profile obtained from a one nanosecond simulation of streptavidin-biotin rupture showing a series of subsequent force peaks most of these can be related to the rupture of individual microscopic interactions such as hydrogen bonds (bold dashed lines indicate their time of rupture) or water bridges (thin dashed lines).

Figure 18.14 NHS-SS-PEG4-biotin can be used to label a primary antibody molecule that has specificity for a protein or interest. Incubation of the biotinylated antibody with a sample, such as a cell lysate, allows the antibody to bind to its target. Capture of the antibody-antigen complex on an immobilized streptavidin reagent effectively isolates the targeted protein from the other proteins in the sample. The disulfide linkage in the spacer arm of the biotin tag permits elution of the immune complex from the streptavidin support using DTT and without using the strong denaturing condition typically required to break the streptavidin-biotin interaction.

Shuvaev, V., Dziubla, T. D., Rainer, W., and Muzykantov, V., Streptavidin-biotin cross-linking of therapeutic enzymes with carrier antibodies nanoconjugates for protection against endothelial oxidative stress, in Methods in Molecular biology , (C. Niemeyer, ed.), Humane Press, Louisville, KY (in press). 

Several qualitative and quantitative immunochemical methods for CAP analysis in biological matrices of animal origin have been described [101,102, 104,105] (see Table 3). Van de Water et al. [ 102] described an ELISA that detected CAP in swine muscle tissue with an IC50 value of 3 ng mL1. This immunoassay was improved and subsequently optimized incorporating the streptavidin-biotin amplification system. There are also several commercially available test kits (see Table 4). RIDASCREEN is a competitive enzyme immunoassay for the quantitative analysis of CAP residues in milk, eggs, and meat in a microtiter plate. The measurement is made photometrically, obtaining a LOD of 100 ng L 1 in meat and eggs and 150 ng L 1 in milk. The test has been also applied to the analysis of tetracyclines. 

All detection systems can be employed following HER and noticeable improvement of most antigen detection will be observed. However, we recommend using sensitive detection systems, such as streptavidin-biotin-peroxidase methods when antigen density is low. 

Musiani, M., Zerbini, M., Plazzi, M., Gentilomi, G., and LaPlaca, M. (1988) Immunocytochemical detection of antibodies to Epstein-Barr virus nuclear antigen by a streptavidin-biotin-complex assay. J. Clin. Microbiol. 26, 1005-1008. 

Bonnard, C., Papermaster, D. S., and Kraehenbuhl, J.-P. (1984) The streptavidin-biotin bridge technique application in light and electron microscope immunocy-tochemistry, in Immunolabelling for Electron Microscopy (Polak, J. M. and Varndell, I. M., eds.), Elsevier, New York, pp. 95-111. 

In the following, protocols for alkaline phosphatase-coupled, peroxidase-coupled antibodies and for the streptavidin-biotin system are given [Jackson and Blythe, 1993 Ormerod and hnrie, 1989]. 

If a biotinylated secondary antibody is used for the streptavidin-biotin method the following protocol is suitable for frozen sections ... 

Make streptavidin-biotin/horseradish peroxidase complex from 20 pi streptavidin, 20 pi biotinylated horseradish peroxidase, 1 ml TBS (shake well and wait 30 min)... 

Let the fluid drain from the sections and incubate with the streptavidin-biotin/horseradish peroxidase complex for 30-60 min Wash with TBS (20 min, 3 times)... 

The third approach made use of the streptavidin-biotin couple to connect enzymes to hydrophobic polymer tails (Figure 6.11c) [36]. In the first step of the self-assembly, two biotin-functionalized polystyrene chains (90 repeat units, PDI =... 

Apart from concentrating the analyte prior to immunoassay, several other possibilities exist for enhancing the sensitivity of the procedure. These constitute introduction of various amplification systems, such as the peroxidase-antiperoxidase system (110), the streptavidine-biotine system (111, 112), and the substrate amplification system (113, 114), which favor the enhancement of the signal per ligand. 

---