Antibodies to trinitrotoluene (TNT) were encapsulated in optically transparent sol-gel derived silica glasses and retained their ability to bind TNT. Both competitive and displacement immunoassays were successfully performed using sol-gel immobilized antibodies. TNT concentrations on the order of ppm were detected. In competitive immunoassays using the sol-gel immobilized antibodies, a logarithmic decrease in fluorescence signal as a function of TNT concentration was observed, similar to competitive immunoassays performed in solution. When encapsulated in the sol-gel. silica matrix, the antibodies retained their ability to differentiate between TNT and trinitrobenzene (TNB), an analogue. In displacement immunoassays, the rate of displacement was dependent upon pore morphology, with aged gels exhibiting faster rates than that of xerogels. The relative stability of antibodies was better for sol-gel encapsulated antibodies than for antibodies immobilized using surface attachment. After exposure to HCl, methanol, or 60 degrees C, the sol-gel immobilized antibodies experienced essentially no loss in ability to bind TNT whereas the surface immobilized antibodies showed as much as 30% loss in ability to bind TNT.