Avidin using SATA

To purify the SATA-modified (strept)avidin use gel filtration on a desalting column or dialyze against 0.1M sodium phosphate, 0.15M NaCl, pH 7.2, containing lOmM EDTA. At this point, the derivative is stable and may be stored under conditions which favor long-term (strept)avidin activity. 

The following protocol describes the activation of avidin or streptavidin with sulfo-SMCC and its subsequent conjugation with an enzyme modified to contain sulfhydryls using SATA (Chapter 1, Section 4.1). A method for the opposite approach, wherein the enzyme is activated with SMCC and the avidin component is thiolated, is presented immediately after this protocol. This strategy may be the most common approach to forming these conjugates (Fig. 363). In addition, since there are enzymes commercially available that are preactivated with SMCC (Pierce), their use may be the easiest solution. 

SATA has been used to form conjugates with avidin or steptavidin with excellent retention of activity (Chapter 23, Section 3.1). It also has been used in the formation of a therapeutically useful toxin conjugate with recombinant CD4 (Ghetie et al., 1990), to study syntaxin proteins (Amessou et al., 2007), to prepare bispecific antibodies (Lindorfer et al., 2001), and to make a unique polylysine conjugate as a vehicle for drug delivery (Sakharov et al., 2001). 

SATA has been used to form conjugates with avidin or steptavidin with excellent retention of activity (Chapter 13, Section 3.1). It has been used in the formation of a therapeutically useful toxin conjugate with recombinant CD4 (Ghetie etal., 1990A). 

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