Monolayers of di-6(A),6(B)-deoxy-6-(4-pyridylmethyl)amino-gamma-cyclodextrin (gamma-CD-(py)(2)) have been formed on polycrystalline platinum electrodes and investigated using electrochemical and surface-enhanced Raman spectroscopy (SERS). The behavior of self-assembled monolayers of (gamma-CD-(py)(2)) alone, (gamma-CD-(py)(2)) backfilled with 1-nonanethiol, and 1-nonanethiol are reported. The potential dependence of the capacitance indicates that the film capacitance is higher for the backfilled CD layers than for 1-nonanethiol layers, most likely due to ion flux through the CD cavity. SERS spectra of the backfilled layer exhibit features associated with both pyridine-functionalized CD and alkane moieties. Investigations using [Fe(CN)(6)](4-) as a solution-phase probe indicate that the backfilled CD-alkane thiol layer exhibits enhanced blocking properties compared to gamma-CD-(py)(2) films alone. Complete blocking was achieved by a combination of backfilling and insertion of a high-affinity guest 1-adamantylamine into the cavity. Significantly, an electroactive guest with high affinity for gamma-CD, [Co(biptpy)(2)](2+), does not exhibit a redox response at the gamma-CD-(py)(2) layer but molecular recognition is turned on by backfilling the CD layer with 1-nonanethiol molecules. This switching on of the electrochemical activity suggests that the CD hosts are initially inaccessible but reorientate upon backfilling, exposing the CD opening to solution and permitting a supramolecular host-guest complex to form. The binding of [Co(biptpy)(2)](2+) to gamma-CD in the backfilled monolayer depends on the bulk concentration of guest and is modeled by the Langmuir isotherm, yielding an association constant for the Co2+ state of 1.45 +/- 0.46 x 10(5) M-1 and a limiting surface coverage 1.49 +/- 0.25 x 10(-11) mol cm(-2). The surface coverage of the divalent state is higher than the trivalent state, reflecting the dynamic nature of the inclusion.