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Biotin Binding Assay (HABA) (PDF)
Biotin Binding: The biotin-binding activity of streptavidin is determined using a modification of the dye-binding assay of Green (1970). One unit will bind one microgram of d-biotin at pH 7.0.
(106mg/g)(A-B)MV 141(A-B) Units/mg = ------------------------- = ----------------- E(Cv) C
Suggestions for Use: Bayer (1989) reports that streptavidin may form aggregates under certain conditions. Streptavidin is highly soluble under alkaline conditions (pH > 8.5). Streptavidin is often supplied lyophilized. Under these conditions there is a tendency for the material to aggregate if it is redissolved in water or other low ionic strength buffers at neutral or acidic pH. As a convenience to customers, Streptavidin has been lyophilized from a dilute sodium chloride solution at mildly alkaline pH. This material is readily soluble in water. The activity of the material recovered after reconstitution under these conditions is undiminished. We recommend dissolving streptavidin in de-ionized water or, preferably, 1.0 mM sodium bicarbonate buffer (pH 9) at twice the desired final protein concentration. The protein may then be diluted with an equal volume of 2x buffer to produce a stock solution. Upon standing some turbidity may develop in certain buffers. Centrifugation will usually yield a clear solution with negligible loss of streptavidin.
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