Antibodies reactions with sodium

Sodium azide is a powerful inhibitor of HRP, but sodium azide can be used with alkaline phosphatase conjugated antibodies without harmful effects. In addition, tap water or water deionized with polystyrene resins may inactivate the enzyme conjugate. Only use distilled, deionized water Tween-20 may interfere with some antibody-antibody reactions or may wash the protein of interest off the membrane. Tween-20 may be left out of the washes, but this may result in increased background... 

This step is optional. The biotin hydrazone-antibody conjugate formed in this reaction (steps 6 and 7) is considered by some researchers to be relatively unstable. To reduce the conjugate to a more stable, substituted hydrazide, treat the conjugate with sodium cyanoborohydride at a final concentration of 5 mA/by adding a 1/20 vol of a 100-mM stock solution. Incubate for 2 h at 4°C (see Note 5). 

The bound peroxidase is visualized by reaction with a filtered solution of 0.05% DAB and 0.0005% hydrogen peroxide in PBS. After rinsing with 0.15 M sodium phosphate buffer (pH 7.4), the sections are postfixed with 0.25% osmium tetroxide in the same buffer for 1 hr and counterstained with 1% uranyl acetate in deionized water. The sections are dehydrated and flat-embedded in Epon according to standard procedures (Hayat, 2000a). Controls are processed as above, except that incubation in the absence of the primary antibody is carried out with PBS containing 2% normal serum and 0.015% Triton X-100. 

Model studies for the syntheses of phosphonate analogues of sphingomyelin and ceramide 1-phosphate from pentaco-ordinate phospholene 112 have been reported. Compound 112 undergoes ring opening on reaction with dialkyl azodicarboxylates to give 113 which reacts with sodium borohydride stereoselec-tively to give predominantly trans- 4 (Scheme 6). The preparation has been described, via phosphorus chloride methods, of the potential haptens 115 for the production of phospholipase A2-like catalytic antibodies. ... 

In an alternative approach, Nakayama and Schultz [25] have successfully achieved the enantiofacial reduction of prochiral ketones. By utilizing the phosphonate hapten 15 catalytic antibodies were elicited which catalyze a highly stereospecific reduction of ketone 16 with sodium cyanoborohydride as a cofactor (Scheme 4). The most active antibody, A5, was found to have a pH optimum at acidic pH, consequently the reductions were performed in aqueous buffer at pH 5.0. The reaction was followed for multiple turnovers (>25) without any decrease in activity or stereoselectivity highlighting the utility of this catalytic system. 

Reduction of periodate-oxidized SVI (14, SVI-IO4) with sodium boro-hydride gave a product (16, SVI-IO4-BH4) which precipitated about three-fifths of the antibody nitrogen precipitated by SVI-IO4 from anti-Pn VI horse serum, and about half that precipitated in the homologous reaction. From cross-reactions of guar gum in anti Pn VI absorbed with SVI-I04 and SVI-IO4-BH4, Rebers and coworkers " showed that some of the antibodies in anti-Pn VI, which precipitated with polysaccharides containing multiple end-groups of D-galactose, also precipitated with SVI-IO4, and they thus obtained additional evidence for the proposed, cyclic structure (IS). 

Similar aza-Diels-Alder reactions of Danishefsky s diene with imines or aldehydes and amines in water took place smoothly under neutral conditions in the presence of a catalytic amount of an alkaline salt such as sodium triflate or sodium tetraphenylborate to afford dihydro-4-pyridones in high yields (Eq. 12.49).117 Antibodies have also been found to catalyze hetero-Diels-Alder reactions.118... 

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