One of the main themes in Sanctuary explores the societal consequences arising from the waning influence of tradition on Southerners in the early 1900s.

Temple Drake, daughter of a wealthy judge, and Popeye, an orphaned child of an impoverished, abandoned mother and a syphilitic wanderer, share a similar flaw. They lack any spiritual traditions handed down from their parents or culture. For them, identity isn't something that naturally grows from internalized values; instead, it is a collection of possible roles they might assume or discard based on their whims. Empty at their core, they search for meaning through romantic relationships, only to end up causing or committing murders.

Horace Benbow, the main character, tries to uphold the Christian tradition he still believes in. To maintain his faith, he clings to several illusions, the most important being the belief that God, through His providence, ensures some form of justice in the world. He believes that by following "the rules," he will succeed and validate those rules. However, almost no one else follows any rules, except for the rule of not getting caught breaking the law or the rule of appearing to live by traditional values. The young people Horace meets and the legal system he deals with—including lawyers, judges, and senators—seem to primarily follow the first rule of avoiding capture. His sister and "the respectable women of Jefferson" adhere to the second rule, which dictates never helping someone in trouble if they are considered disreputable.

The people in trouble are a family of bootleggers, viewed as disreputable figures who serve the community by supplying high-quality illegal liquor but must be shunned when discovered.

This central theme is apparent at nearly every societal level and in every character. Without a sustaining tradition, society becomes barren, and life turns into a nightmare. The only potential challenge to this horror comes from the faint voices of a few characters who remain anchored in tradition, dream of spiritual fulfillment, or are committed to love. Horace's elderly aunt, secure in tradition, seems out of place in the modern world. A young boy dreaming of a better future falls under the corrupt influence of Senator Clarence Snopes. The one woman committed to love is Lee's common-law wife, Ruby Lamar; her baby is portrayed as a counterpart to Popeye.

Horace Benbow, the main character, strives to maintain the Christian values he still holds dear. To keep his faith, he clings to several illusions, the most important being the belief that God, through divine providence, ensures some form of justice in the world. He believes that by following "the rules," he will succeed and thereby affirm those rules. However, nearly everyone else seems to ignore any rules, except for the one about not getting caught breaking the law or the one about keeping up appearances of adhering to traditional values. The young people Horace meets and the legal system he deals with — including lawyers, judges, and politicians — mainly abide by the first rule of avoiding capture. His sister and "the respectable women of Jefferson" stick to the second rule, which states that one should never assist someone in trouble if that person has a bad reputation.

Popeye and Temple cross paths by chance at the hideout of the Goodwins, who are bootleggers. Attracted to Temple, Popeye eventually assaults her with a corn cob due to his impotence and then kidnaps her, taking her to a brothel in Memphis. There, he arranges for her to be with another man, trying to live through their encounters. When Temple starts enjoying this too much, Popeye separates them. When...

In Sanctuary, contemporary American society seems headed for self-destruction. The voices offering salvation are barely audible. People with good intentions find themselves trapped by illusions. Power is held by empty individuals who temporarily overlook the obvious despair and emptiness surrounding them. When they, like Popeye, recognize the void, they will permit themselves to die. Until that moment, they will continue to kill, both directly and indirectly, as Temple and numerous other characters do.