IEEE International Conference on “Nanomaterials: Applications & Properties” (NAP-2020) Sumy, Ukraine, 9-13 Nov. 2020 XXX-X-XXXX-XXXX-X/XX/$XX.00 ©20XX IEEE IDNUM-1 Sequential magnetic mapping of bacteria loaded with Pd-Fe nanoparticles James Claxton Department of Physics University of Oslo Oslo, Norway j.b.claxton@fys.uio.no Dirk Linke Department of Biosciences University of Oslo Oslo, Norway dirk.linke@ibv.uio.no Nadeem Joudeh Department of Biosciences University of Oslo Oslo, Norway nadeem.joudeh@ibv.uio.no Pavlo Mikheenko Department of Physics University of Oslo Oslo, Norway pavlo.mikheenko@fys.uio.no Anja Røyne Department of Physics University of Oslo Oslo, Norway anja.royne@fys.uio.no Abstract— Magnetic nanoparticles are of widespread use in nanotechnology. One of the most unusual are magnetic palladium nanoparticles that combine magnetism with high catalytic activity. These nanoparticles could be obtained biologically by exposing bacteria to a palladium salt. Due to their small size and weak magnetism, however, it is challenging to measure their magnetic properties. One of the solutions to enhance their magnetism is to incorporate a small amount of iron atoms into them. After this procedure, the nanoparticles together with bacteria can be embedded in resin and characterized by the technique of magnetic force microscopy. This technique allows imaging cross-sections of the bacteria with nanoparticles, but cannot give information from the depth of the sample. Here we report on an approach partially solving this problem. Its novelty lies in measurements of consecutive thin slices of resin, which allows mapping cross-sections of individual bacteria and different parts of the material surrounding the same bacterium. An interesting observed feature is the formation of magnetic chains of nanoparticles outside of the bacteria. Keywords— Nanoparticles, magnetic mapping, palladium, iron, bacteria, magnetic force microscopy I. INTRODUCTION Magnetic nanoparticles are becoming indispensable tools in nanotechnology. Their applications range from magnetic recording media [1] to delivery of drugs [2] and treatment of cancer [3]. Special attention is attracted to palladium (Pd) nanoparticles that become magnetic when their size is in the range of a few nanometers [4,5]. Combined with excellent catalytic properties, these nanoparticles are especially efficient in cancer treatment [6]. A biological method for producing Pd nanoparticles in very large amounts is to introduce bacteria to a Pd salt solution [7]. In their exchange with the environment, bacteria can supply electrons for redox reactions, and efficiently reduce Pd salts from solution due to very high redox potential of the latter [8,9]. Adding iron (Fe) salt to the solution allows obtaining Pd particles that incorporate a small amount of Fe, which strongly increases their magnetism [10]. It could be straightforward to use these Pd-Fe nanoparticles for applications, but it is not easy to measure their magnetic properties. Here a technique is reported allowing doing this with the help of magnetic force microscopy (MFM). II. EXPERIMENTAL A. Magnetic force microscopy Magnetic force microscopy (MFM) [11-13] is a technique allowing mapping magnetic properties of a sample with a magnetic tip, which is scanned above its surface. In order to distinguish between magnetic and Van der Waals forces, which are acting on the tip at small distances, a two- pass scanning technique is used [11,12]. In the first pass, topography close to the surface is mapped. In the next scan, the probe is moved along a path following the measured topography, but at a larger height, so that the probe-sample distance is kept constant. If the height is large enough, Van der Waals forces become weak, and the pure magnetic response can be measured. According to [11,13,14], shift in the phase of oscillations, if AC mode is used, is proportional to the gradient of force acting on the tip. The measurements were done using JPK NanoWizard 4.0 in AC mode at a frequency of about 74 kHz in an applied field of 0.58 T, created by a permanent magnet within the sample holder. The probes used were manufactured by Nanosensors, model type PPP-MFMR-10, with a tip radius of approximately 50 nm. B. Production and preparation of samples A single colony of Escherichia coli BW25113 strain was inoculated into 10 ml lysogeny broth (LB) medium in a test tube overnight at 37  C while shaking it at 200 rpm. On the next day, 1 ml of this medium was used to inoculate 49 ml fresh LB medium in a 250 ml flask. The flask was also incubated at 37  C while shaking at the same speed until the optical density (O.D.600) reached 0.5. The medium was transferred to a 50 ml falcon tube and centrifuged at 4250 g for 10 mins. The supernatant was removed, and the pellet was resuspended in 10 ml 20 mM pH 7 3-(N- morpholino)propanesulfonic acid (MOPS) buffer. This washing step was done two more times, except for the last round the pellet was resuspended in 8 ml 20 mM pH 7 MOPS buffer. 1 ml of this suspension was transferred to a 1.5-ml Eppendorf tube. 1 mM of sodium tetrachloropalladate (Na2Cl4Pd) and 1 mM of iron III chloride (FeCl3) (both dissolved in 0.01M nitric acid) were added to the tube, shaken well by hand and incubated for 1 hour. Then, 10 mM