Prevent users from uploading the same file twice using EnableDuplicateCheck="true".
Use DuplicateHashAlgorithm to specify the hashing method for content-based detection.
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<au:AjaxFileUpload ID="Uploader1" runat="server"
AllowMultiple="true"
EnableDuplicateCheck="true"
DuplicateHashAlgorithm="sha256"
ButtonText="Select Files"
AutoUpload="true"
ShowProgress="true" />
Try selecting the same file twice to see it detected as a duplicate.