Circular-difference response of second-harmonic generation from chiral surfaces is shown to be a sensitive probe of chiral biological samples. The sensitivity of this nonlinear effect is several orders of magnitude higher than that of linear circular dichroism and is demonstrated using samples of bacteriorhodopsin. This nonlinear optical activity probes molecular chirality on a more general level than linear techniques. Surface specifity of the novel approach can be of considerable importance in the study of biological membranes. Hence, this process has the potential of becoming a new chiro-optical tool in the fields of analytical (bio)chemistry, surface chemistry, and cell biology.