RESEARCH COMMUNICATIONS CURRENT SCIENCE, VOL. 110, NO. 12, 25 JUNE 2016 2297 *For correspondence. (e-mail: ys_rajput@rediffmail.com) A method for synthesis of gold nanoparticles using 1-amino-2- naphthol-4-sulphonic acid as reducing agent Pooja Rawat 1 , Y. S. Rajput 1, *, Mahesh K. Bharti 1 and Rajan Sharma 2 1 Animal Biochemistry Division, and 2 Dairy Chemistry Division, ICAR-National Dairy Research Institute, Karnal 132 001, India The present communication describes the synthesis of gold nanoparticles (AuNPs) using 1-amino-2- naphthol-4-sulphonic acid (ANSA) with enhanced sta- bility. The method of preparation is similar to the conventional method using sodium citrate as reducing agent. Briefly, ANSA was quickly added to a boiled solution of hydrogen tetrachloroaurate under stirring, resulting in the synthesis of deep red-coloured AuNPs. The AuNPs prepared by ANSA (AuNPs–ANSA) exhibit twofold increase in stability towards sodium chloride over those prepared by sodium citrate. AuNPs–ANSA are 35.1 nm in size and particles has various shapes, viz. hexagonal, pentagonal, spherical, etc. AuNPs–ANSA can be functionalized with immu- noglobulins and functionalized nanoparticles exhibit flow properties in lateral-flow assay. Keywords: Gold nanoparticles, immunoglobulins, lateral flow assay, reducing agent. GOLD nanoparticles (AuNPs) offer unique physical, elec- tronic, magnetic, thermal, optical and biomedical proper- ties. High absorption coefficient, large surface area, ease of conjugation to various biomolecules and excellent biocompatibility make them suitable for several applica- tions. These particles have found applications in sensing, diagnostics, treatment and catalysis 1–12 . Conventionally, AuNPs were prepared by reduction of tetrachloroauric acid with sodium citrate in aqueous environment 13–15 . The reduction essentially requires addition of sodium citrate to boiled solution of tetrachloroauric acid under constant stirring. Citrate ions provide negative surface charge to AuNPs by adhering to their surface, and this results in the stabilization of these particles. A number of other reduc- ing and stabilizing agents such as EDTA, amines, glyc- erol, appin and hydroquinone have been employed for gold colloid preparation 16–20 . These reducing agents result in 10–50 nm sized AuNPs. Smaller sized (1.2–2.8 nm) AuNPs can also be prepared in organic phase by reduc- tion of chloro (triethylphosphine) gold (Et 3 PAuCl) with 9-borabicyco[3,3,1]nonane (9-BBN) 21 . Binding of electron-donating end group of a ligand such as poly (allylamine) hydrochloride or triphenylphosphine to the inorganic nanoparticle core has also been reported 17,21–23 . AuNPs are destabilized by cations, and this restricts their application within threshold limits of ion concentration. The reducing agent used in the present study is 1-amino- 2-naphthol-4-sulphonic acid (ANSA), which may reduce tetrachloroauric acid as well as impart negative charge on the colloid surface (Figure 1). The high ionic stability of nanoparticles prepared by this method offers added advantage in comparison to gold colloids prepared by citrate reduction method. Size appropriateness is essential for in vivo applications. Particles smaller than 10 nm are likely to diffuse non-specifically into non-target tissues and organs, whereas particles larger than 100 nm are likely to get trapped in the liver and lung 24–26 . In comparison to citrate reduction, the method presented here provides nanoparticles of large diameter (35 nm) which is also advantageous in lateral flow techniques due to their better visibility without compromising on flow property. Hydrogen tetrachloroaurate (III) (HAuCl 4 ) and sodium citrate tribasic dihydrate were obtained from Sigma- Aldrich (St. Louis, MO, USA). ANSA was obtained from Sisco Research Laboratories Pvt Ltd, Mumbai, India. Rabbit immunoglobulin G (IgG) and goat anti-rabbit IgG (affinity-purified) were from Merck Millipore (Ben- galuru). All other reagents were of analytical grade. The materials required for the fabrication of lateral flow strip (backing card glued with nitrocellulose membrane in the mid portion, sample pad and adsorbent pad) were pro- cured from Advanced Microdevices Pvt Ltd (Ambala). All the glassware used for nanoparticles synthesis was treated overnight with aqua regia, rinsed with generous amounts of double-distilled water and then MilliQ water before use. The water employed in the study was MilliQ (18.2 M). Particle size analysis was performed using Malvern Zetasizer ZS90, UK. Spectral analysis was per- formed using a UV-visible spectrophotometer (Shimadzu Analytical Pvt Ltd, Mumbai). TEM images were acquired using a transmission electron microscope (FEI TECNAI G2 F20, Oregon, USA). Figure 1. Depiction of synthesis and stabilization of gold nanoparti- cles (AuNPs) by 1-amino-2-naphthol-4-sulphonic acid (ANSA) from HAuCl4.