We explored the sensitivity and selectivity of gold nanoparticles (AuNPs) treated paper as a generic SERS diagnostic platform to identify and quantify low concentrations of a specific (bio)analyte in aqueous solutions. The effects of gold nanoparticles (AuNPs) concentration on their adsorption and aggregation states on paper were explored. The surface coverage of AuNPs on paper scaled linearly with their concentration profile in solutions. The SERS performances of the AuNPs-treated papers were evaluated with a model Raman molecule, 4-aminothiophenol (4-ATP), and their SERS intensities increased linearly with the density of AuNPs on paper. To increase the SERS sensitivity, the retention and aggregation state of nanoparticles on paper was controlled by pre-treating paper with a series of cationic polyacrylamide (CPAM) solutions. The CPAM pre-treated paper produced a more uniform distribution of AuNPs compared to untreated paper. Higher surface coverage and aggregation of AuNPs on paper were favoured by CPAM solutions of higher concentration, charge density and molecular weight. The optimized AuNPs-CPAM paper showed a higher sensitivity and Raman enhancement factor (EF), which was almost an order of magnitude higher than the untreated AuNPs paper. After the SERS sensitivity towards the detection of model Raman molecule (4-ATP) was proven, the SERS selectivity of AuNPs paper was demonstrated by functionalizing the AuNPs with a model biomolecule platform consisting of biotin/streptavidin assemblies for the detection of antibody-antigen binding. The modification of antibody local structure due to the interaction with antigen was detected. Evidence of antigen binding was elucidated from the SERS spectra, confirming the presence of antigen. Reproducible spectra features were observed for the functionalized AuNP papers which were exposed to different concentration of antigen; the spectra intensity increased as a function of antigen concentration. The sensitivity and selectivity of AuNPs paper substrates as a low-cost and generic SERS platform for bio-diagnostic application was demonstrated.