Nonlinear laser wave-mixing spectroscopy is presented as an ultrasensitive detection method for chemical and biological agents in thin-film and liquid-phase samples. Wave mixing is an unusually sensitive absorption-based detection method that offers inherent advantages including excellent sensitivity, small sample requirements, short optical path length, high spatial resolution and excellent standoff detection capability. Wave mixing offers excellent optical absorption detection sensitivity even when using thin samples (<0.1 mm), and hence, it is inherently suitable for interfacing to microarrays, microfluidics, and capillary electrophoresis. Laser excitation wavelengths can be tuned to detect multiple chem/bio agents in their native form. Since the wave-mixing signal is a coherent laser-like beam with its own propagation direction, it offers excellent S/N and allows remote standoff detection capability. Laser wave mixing is presented as a way to natively detect small biomolecules and nitroaromatic explosives at zeptomole mass detection limits. The absorbance of select neurotransmitters and nitroaromatic explosives at ultraviolet wavelengths make their detection possible using a compact pulsed UV-laser. The detection of these analytes is successfully accomplished both on UV-transparent surfaces and by using capillary electrophoresis separation. Wave mixing is used for the ultrasensitive detection of cellular proteins and antibodies using fluorescent and non-fluorescent labels. Using sodium dodecyl sulfate capillary gel electrophoresis, proteins can be separated by their size using an appropriate sieving matrix. Newly developed methods employ wave-mixing and capillary electrophoresis for the zeptomole mass detection of cellular proteins and antibodies without the need for time consuming capillary preparation steps. The HIV-1 p24 capsid protein has been detected using laser wave-mixing spectroscopy and capillary electrophoresis using chromophore and fluorophore labels. Size-based capillary electrophoresis separation has also been used to analyze the products of a p24 antibody-antigen reaction. These studies show the potential of wave mixing to be used to create field-deployable and relatively inexpensive HIV viral load screens in resource-limited settings.