*For correspondence. E-mail: saloutim@yahoo.com; Tel. & Fax: +98-241- 4224024 Rasoul Shokri 1 , Mojtaba Salouti 2 * , and Rahim Sorouri Zanjani 3 1 Department of Microbiology, Sciences and Research Branch, Islamic Azad University, Fars, Iran 2 Biology Research Center, Zanjan Branch, Islamic Azad University, Zanjan, Iran 3 Zanjan University of Medical Sciences, Zanjan, Iran (Received Sep 22, 2014 / Revised Nov 17, 2014 / Accepted Nov 18, 2014) Journal of Microbiology (2015) Vol. 53, No. 2, pp. 116–121 Copyright  2015, The Microbiological Society of Korea DOI 10.1007/s12275-015-4519-4 Anti protein A antibody-gold nanorods conjugate: a targeting agent for selective killing of methicillin resistant Staphylococcus aureus using photothermal therapy method The high prevalence of methicillin resistant Staphylococcus aureus (MRSA) and developing resistance to antibiotics re- quires new approaches for treatment of infectious diseases due to this bacterium. In this study, we developed a target- ing agent for selective killing of MRSA using photothermal therapy method based on anti protein A antibody and gold nanorods (GNRs). Polystyrene sulfonate (PSS) coated GNRs were conjugated with anti protein A antibody. The FT-IR and UV-vis analyses approved the formation of anti protein A antibody-gold nanorods conjugate. In vitro study of pho- tothermal therapy showed 82% reduction in the MRSA cells viability which was significantly greater than the ablation ef- fect of free GNRs and laser alone. Significant accumulation of anti protein A antibody-GNRs in the infected muscle in comparison with normal muscle approved the targeting abi- lity of new agent. In vivo study of photothermal therapy re- sulted in a significant reduction (73%) in the bacterial cells viability in the infected mouse model. These results demon- strated the ability of anti protein A antibody-GNRs conjugate in combination with NIR laser energy for selective killing of MRSA in mouse model. Keywords: Methicillin Resistant Staphylococcus aureus, pho- tothermal therapy, gold nanorods, anti protein A antibody Introduction Methicillin-resistant Staphylococcus aureus (MRSA), a Gram- positive spherical cells usually arranged in grape-like irregu- lar clusters, is responsible for several difficult-to-treat infec- tious diseases in humans such as bacteremia, endocarditis, meningitis, impetigo, eczema, and folliculitis (Huang, 2006; Fernandes et al., 2012; Galanzha et al., 2012; Turos et al., 2012). Unfortunately, antiseptics such as hexachlorophane, triclosan, chlorhexidine, and povidoneiodine are not fully effective against MRSA and there are some evidences of re- sistance developing to these agents (Zharov et al., 2006; Zol- faghari et al., 2009). Given that MRSA has evolved mecha- nisms of resistance for most commercially produced anti- biotics, it is necessary to develop novel strategies not rely on traditional therapeutic regimes (Turos et al., 2007; Huang et al., 2009). One such strategy involves photothermal therapy method using the rod shape of gold nanoparticles to destroy bacteria (El-Sayed et al., 2006; Chen et al., 2008; Pissuwan et al., 2008). Here we describe the approach involving physical damage to MRSA using the combination of NIR laser energy and absorbing nanoparticles selectively attached to the bacte- rial cells. When gold nanoparticles are irradiated, they absorb energy, which is quickly transferred through non radiative relaxation into the heat and accompanied effects, and even- tually leads to irreparable damages. Gold nanorods are pho- tostable and nontoxic material that manifest a low level of toxicity when introduced into the biological systems and are easily conjugated to the antibodies or proteins (Panyala et al., 2009). In addition, their maximum absorption can be shifted to near-infrared radiation (NIR) ranging from 700 to 900 nm, where the minimum absorption of light occurs in most human tissues, allowing deeper penetration of laser radiation into the infected tissues. (Grace and Pandian, 2007; Huff et al., 2007). Therefore, gold nanorods can be useful as a photothermal therapy agent since their use would mini- mize the risk of damages to healthy cells (Mohanpuria et al., 2008; Pissuwan et al., 2008). To date, laser energy in combination with targeted GNRs has been successfully used in killing bacterial cells in vitro, but selective ablation of bacterial infection foci using pho- tothermal therapy method has not been performed in vivo. Norman et al. (2008) used gold nanorods conjugated with primary antibodies for killing of pathogenic Gram-negative bacterium, Pseudomonas aeruginosa, in vitro. They found a significant reduction in bacterial cells viability following ex- posure to near-infrared laser. In this study, we performed the experiments in which GNRs were covalently conjugated with anti protein A antibody to produce a nanodrug for selective killing of MRSA using photothermal therapy method in vitro and in vivo. Anti protein A antibody is a monoclonal anti- body against the specific product of Staphylococcus aureus, protein A, one of the major surface-clustered of proteins that is linked to the peptidoglycan portion of the bacterial cell walls. About 99% of S. aureus strains express protein A on their cell walls (Huang, 2006). So, it seems a good target for selective killg of MRSA.