Immunoglobulins incorporation

Probe for neurological function Immunoglobulin incorporation using staphylococcal protein A Mylein basic protein... 

An electrode in which an antibody or an antigen/hapten is incorporated in the sensing element is termed an immunoelectrode . The potential response of the immuno-electrode is based on an immunochemical reaction between the sensing element of the electrode and antibody or antigen/hapten in the sample solution. One example of such an electrode is the polymer membrane electrode shown in Fig. 7. The selective response of this electrode to specific immunoglobulins is based on the interaction between antibody in solution and an antigen-ionophore complex in the membrane ... 

With mixing, add a quantity of the sulfo-NHS-biotin solution to the protein solution to obtain a 12- to 20-fold molar excess of biotinylation reagent over the quantity of protein present. For instance, for an immunoglobulin (MW 150,000) at a concentration of 10 mg/ml, 20 pi of a sulfo-NHS-biotin solution (containing 8 X 10-4 mmol) should be added per ml of antibody solution to obtain a 12-fold molar excess. For more dilute protein solutions (i.e., 1-2 mg/ml), increased amounts of biotinylation reagent may be required (i.e., 20-fold molar excess or more) to obtain similar incorporation yields as when using more concentrated protein solutions. 

Add 0.3 mg of sulfo-NHS-SS-biotin (Thermo Fisher) to each ml of the antibody solution. To measure out small amounts of the biotinylation reagent, it may be first dissolved in water at a concentration of at least 1 mg/ml. Immediately transfer the appropriate amount to the antibody solution. This level of sulfo-NHS-SS-biotin addition represents about an 8-fold molar excess over the amount of antibody present. This should result in a molar incorporation of approximately 2-4 biotins per immunoglobulin molecule. 

Jackson et al. (1988) were the first to employ the immunogold-silver staining (IGSS) method in combination with microwave heating. They completed within minutes the incubations in primary and secondary antibodies for detecting immunoglobulins in paraffin sections of human tonsil, van de Kant et al. (1990) applied the same method, except that resin instead of paraffin sections were used to detect bromodeoxyuridine incorporated in cells of the mouse testis. Tissue morphology is preserved better in a resin than in paraffin. The former also allows the use of thinner sections. 

Nucleic acid probes are commonly labeled by enzymatic incorporation of radiolabeled nucleotides or enzymatic addition of radiolabeled phosphate groups to the nucleic acid chain. Proteins, in particular immunoglobulins, are labeled commonly by direct... 

Acceptable bridging molecule systems have been developed which have also simplified the utilization of different detection systems. To illustrate this point, a researcher who has developed a unique monoclonal antibody (a primary antibody) in the mouse may select from a variety of commercially available products consisting of different detection systems (e.g. fluorescein, alkaline phosphatase, colloidal gold) attached to an immunoglobulin that will specifically bind to mouse antibodies (a secondary antibody). In this way the researcher may readily obtain and test a number of detection methods for visualizing target-probe interactions without having to directly label the monoclonal antibody probe. For nucleic acid probes, which in themselves are not readily immunodetectable, it is useful to incorporate or attach detectable moieties to the nucleotides. 

Linder V, Verpoorte E, de Rooij NF, Sigrist H, Thormann W. Application of surface biopassivated disposable poly(dimethylsiloxane)/glass chips to a heterogeneous competitive human serum immunoglobulin G immunoassay with incorporated internal standard. Electrophoresis 2002 23 740-749. 

The labeling of BiP in mouse myeloma B cells by [ H]adenosine, suggestive of ADP-ribosylation, has been shown to be a modification found on BiP free of immunoglobulin heavy chains, but not on BiP bound to heavy chains (Hendershot et al., 1988). Additionally, the [ H]adenosine labeling parallels BiP phosphorylation in vivo conditions that increase BiP production decrease BiP phosphorylation and [ HJadenosine incorporation, suggesting that these two posttranslational modifications might modulate BiP activity in concert in some manner. 

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