Selection of aptamers against Lactoferrin based on silver enhanced and fluorescence-activated cell sorting

We report a novel method for efficiently screening aptamers from a complex ssDNA library based on silver decahedral nanoparticles (AgNP) and fluorescence activated cell sorting (FACS). In this method, target protein (lactoferrin) and negative proteins (alpha-lactalbumin, beta-lactoglobulin, bovine serum albumin, casein) were respectively immobilized on polystyrene microspheres (PS) to form PSLac, PS alpha-Lac, PS beta-Lac, PSBSA and PSCas. PSLac was firstly interacted with Cy5 labeled library (Lib), then hybridized with Cy5 modified silver decahedral nanoparticles (AgNPCy5) to form PSLac/Lib/AgNPCy5 conjugates. FACS was used to separate and collect PSLac/Lib/AgNPCy5 conjugates from complicated complex. AgNP was used to increase the fluorescence intensity in the selecting process and choose non-self-hybridization of Lib. Six aptamers (Ylac1, Ylac4, Ylac5, Ylac6, Ylac8 and Ylac9) were obtained after five-round of selection. These aptamers showed good specificity towards lactoferrin in the presence of negative proteins. The equilibrium dissociation constants (K-d) of six aptamers were calculated and all were in the nanomolar range. In a word, AgNP-FACS SELEX (AgFACS-SELEX) is a rapid, sensitive and highly efficient method for screening aptamers.