Intravascular or blood pool contrast agents represent a distinct class of contrast media that can remain stable in blood circulation for several hours, enabling better visualization of vasculatures through prolonged imaging. Superparamagnetic iron-oxide nanoparticles (SPION) have emerged as the second generation MRI contrast agents that are widely used to investigate the in vivo vascular parameters such as cerebral blood volume (CBV) and microvascular volume 1, 2. Currently, many of these studies are conducted with Ferumoxytol, a US-FDA approved iron supplement based on carboxymethyl dextran (CMD) covered iron oxide. However, Ferumoxytol is relatively expensive to afford for basic research and not customized for advanced imaging needs. For these reasons, the present study aims to 1) develop a simple, one-pot recipe for producing blood-pool type SPION in-house avoiding costly equipment and complex synthetic strategies and 2) identify some of the key design considerations that can be used to tailor their pharmacokinetics (PK) and relaxivity in vivo in order to comply with specific imaging requirements. Herein, we report the synthesis of three different SPION formulation coated with varying amount of carboxymethyl dextran (CMD), designated as CMD-ION@L, -M and –H respectively in ascending order of CMD content. Using CMD-ION as a model system, we show that relaxivity and PK profile of SPION can be effectively tuned via intelligent manipulation of surface chemistry and that a compromise between the nanomagnetic core and outer polymeric shell is critical to ensure maximal performance in vivo. The synthesis of CMD-SPION has been presented schematically in Figure 1. Size and surface chemistry of CMD-SPION were characterized using transmission electron microscopy (TEM) and X-ray photoelectron spectroscopy respectively. Relaxometry of CMD-ION@L-H was evaluated in vitro with a Siemens Magnetom 7T scanner using a multi spin-echo RARE T1-T2 mapping sequence. PK assessment was done in anesthetized Long Evans rats using a Bruker 9.4T MRI scanner, home-made surface coil and a single shot, single sampled GE-EPI sequence. In order to monitor the clearance of SPION from cerebrovasculature in real time, EPI data was continuously acquired for 4h and animals were intravenously injected with CMD-ION (~10 mg/kg) 15 min after scan onset. For visual stimulation experiments, animals were exposed to visual stimuli (40s OFF-10s ON-40sOFF paradigm repeated 5 times) using 10 Hz laser pulses. Hypercapnia (5% CO2) gas challenge studies were conducted in C57-B6 wild type mouse using a 240s OFF-240sON-240s OFF paradigm. Rat cerebromicroangiography was performed in a Bruker 7T scanner using a 3D RARE sequence 3. Representative 3D view of microvasculature was constructed in Amira using a maximum intensity projection (MIP) utility. CMD coated SPION (5-10 nm) with different surface composition were synthesized using simple, one-pot chemical reactions, easily amenable to commercial scale-up. We observed that surface compositions of as-prepared CMD-IONs were strictly dependent on the molar stoichiometry of the reactants used during the synthesis, which in turn affected the transverse relaxivity (T2) and in vivo stability of the formulations [Fig 1(a-c)]]. As evident from pharmacokinetic imaging, a bolus injection of CMD-ION@L in rats led to almost 50-60% drop in signal intensity in most brain areas (Fig 2a) but the contrast was short-lived (~2h). On the other hand, injection of CMD-ION@H led to very small (only 10-20%) changes in ΔR2* though the contrast lasted over a period greater than 4h. Among all the formulations, CMD-ION@M showed the best performance in terms of both contrast and in vivo stability (elimination half life >8h). To further understand the tissue distribution profile of our home-made SPION, a small group of mice (n=5) was injected with CMD-ION@M (40 mg/kg). Animals were perfused after 7 days post-injection. Prussian blue staining revealed significant iron accumulation in liver and spleen suggesting that CMD-ION@M eventually clears from the intravascular space and metabolizes within macrophages (Fig 2b). Using this optimized formulation, we were able to visualize very robust, target-specific ΔR2* changes (~6%) in rats exposed to visual stimulation (Fig 3a) and almost 10-15% ΔR2* changes in mice challenged with 5% CO2 (Fig 3b). CMD-ION@M also showed excellent performance as a micro-MRA contrast agent, improving the visibility of cerebromicrovessels in vivo (Fig 3c). We have developed a simple, inexpensive method to synthesize high-performance intravascular SPION in house. We were able to tune the relaxivity and PK profile of our home-made SPION via careful surface functionalization control and came up with an optimal formulation that offered very robust and stable contrast for high resolution cerebromicroangiography and steady state CBV functional imaging. Our future studies will focus on developing novel MR-detectable inflammation markers using our home-made iron oxide as the platform material.