<em>In vivo</em> <sup>19</sup>F MRI for Cell Tracking

In vivo 19F MRI allows quantitative cell tracking without the use of ionizing radiation. It is a noninvasive technique that can be applied to humans. ...

Radboud university medical center

We give an overview of the applications and methods of high-resolution anatomical magnetic resonance imaging (MRI) in the study of embryonic and fetal development in animal models. Challenges associated with performing in utero studies are described. Recent in utero images in mouse and in nonhuman primates are presented. Results using magnetic resonance microscopy in fixed mouse embryos and in amphibian embryos in vivo are reviewed. We discuss how studies of pregnancy in animal models aid in the translation of innovative new MRI techniques to clinical applications.

Magnetic resonance imaging of embryonic and fetal development in model systems.

This article describes an in vivo imaging method for visualizing and quantifying a specific cell population. Cells are labeled ex vivo with a perfluoropolyether nanoparticle tracer agent and then detected in vivo using (19)F MRI following cell transfer. (19)F MRI selectively visualizes only the labeled cells with no background, and a conventional (1)H image taken in the same imaging session provides anatomical context. Using the nonobese diabetic mouse, an established model of type 1 diabetes, (19)F MRI data were acquired showing the early homing behavior of diabetogenic T cells to the pancreas. A computational algorithm provided T cell counts in the pancreas. Approximately 2% of the transferred cells homed to the pancreas after 48 hr. The technique allows for both unambiguous detection of labeled cells and quantification directly from the in vivo images. The in vivo quantification and cell trafficking patterns were verified using (19)F spectroscopy and fluorescence microscopy in excised pancreata. The labeling procedure did not affect T-cell migration in vivo. This imaging platform is applicable to many cell types and disease models and can potentially be used for monitoring the trafficking of cellular therapeutics.

Fluorine-19 MRI for visualization and quantification of cell migration in a diabetes model.

We report the design, synthesis, and biological testing of highly stable, nontoxic perfluoropolyether (PFPE) nanoemulsions for dual 19F MRI-fluorescence detection. A linear PFPE polymer was covalently conjugated to common fluorescent dyes (FITC, Alexa647 and BODIPy-TR), mixed with pluronic F68 and linear polyethyleneimine (PEI), and emulsified by microfluidization. Prepared nanoemulsions (<200 nm) were readily taken up by both phagocytic and non-phagocytic cells in vitro after a short (approximately 3 h) co-incubation. Following cell administration in vivo, 19F MRI selectively visualizes cell migration. Exemplary in vivo MRI images are presented of T cells labeled with a dual-mode nanoemulsion in a BALB/c mouse. Fluorescence detection enables fluorescent microscopy and FACS analysis of labeled cells, as demonstrated in several immune cell types including Jurkat cells, primary T cells and dendritic cells. The intracellular fluorescence signal is directly proportional to the 19F NMR signal and can be used to calibrate cell loading in vitro.

Self-delivering nanoemulsions for dual fluorine-19 MRI and fluorescence detection.

Noninvasive methods to image the trafficking of phenotypically defined immune cells are paramount as we attempt to understand adaptive immunity. A (19)F MRI-based methodology for tracking and quantifying cells of a defined phenotype is presented. These methods were applied to a murine inflammation model using antigen-specific T cells. The T cells that were intracellularly labeled ex vivo with a perfluoropolyether (PFPE) nanoemulsion and cells were transferred to a host receiving a localized inoculation of antigen. Longitudinal (19)F MRI over 21 days revealed a dynamic accumulation and clearance of T cells in the lymph node (LN) draining the antigen. The apparent T-cell numbers were calculated in the LN from the time-lapse (19)F MRI data. The effect of in vivo T-cell division on the (19)F MRI cell quantification accuracy was investigated using fluorescence assays. Overall, in vivo cytometry using PFPE labeling and (19)F MRI is broadly applicable to studies of whole-body cell biodistribution.

In vivo cytometry of antigen-specific t cells using 19F MRI.

Cellular therapy, including stem cell transplants and dendritic cell vaccines, is typically monitored for dosage optimization, accurate delivery, and localization using noninvasive imaging, of which magnetic resonance imaging (MRI) is a key modality. (19)F MRI retains the advantages of MRI as an imaging modality, and also allows direct detection of labeled cells for unambiguous identification and quantification, unlike typical metal-based contrast agents. Recent developments in (19)F MRI-based in vivo cell quantification, the existing clinical use of (19)F compounds and current explosive interest in cellular therapeutics have brought (19)F imaging technology closer to clinical application. We review the application of (19)F MRI to cell tracking, discussing intracellular (19)F labels, cell labeling and in vivo quantification, as well as the potential clinical uses of (19)F MRI.

(19)F MRI for quantitative in vivo cell tracking.

Monitoring cell trafficking in vivo noninvasively is critical to improving cellular therapeutics, drug delivery, and understanding disease progression. In vivo imaging, of which magnetic resonance imaging (MRI) is a key modality, is commonly used for such monitoring. (19)F MRI allows extremely specific detection and quantification of cell numbers directly from in vivo image data, longitudinally and without ionizing radiation. We used fluorocarbons previously used in blood substitutes and imaging agents for ultrasound and computed tomography to synthesize monodisperse nanoparticles that are stable at 37 degrees C and can be frozen for storage. These large (19)F labeling compounds are insoluble in aqueous environments and often emulsified, typically forming emulsions unsuitable for long-term storage. Instead, we used a non-toxic polymer already in clinical use, poly(D,L-lactide-co-glycolide), to encapsulate a range of (19)F compounds. These nanoparticles can be customized in terms of content (imaging agent, fluorescent dye, drug), size (200-2000 nm), coating (targeting agent, antibody) and surface charge (-40 to 30 mV). We added a fluorescent dye and antibody to demonstrate the versatility of this modular imaging agent. These nanoparticles are adaptable to multimodal imaging, although here we focused on MRI and fluorescence imaging. Here, we imaged primary human dendritic cells, as used in clinical vaccines.

Customizable, multi-functional fluorocarbon nanoparticles for quantitative in vivo imaging using 19F MRI and optical imaging.

Monitoring of cell therapeutics in vivo is of major importance to estimate its efficacy. Here, we present a novel intracellular label for (19)F magnetic resonance imaging (MRI)-based cell tracking, which allows for noninvasive, longitudinal cell tracking without the use of radioisotopes. A key advantage of (19)F MRI is that it allows for absolute quantification of cell numbers directly from the MRI data. The (19)F label was tested in primary human monocyte-derived dendritic cells. These cells took up label effectively, resulting in a labeling of 1.7 ± 0.1 × 10(13) (19)F atoms per cell, with a viability of 80 ± 6%, without the need for electroporation or transfection agents. This results in a minimum detection sensitivity of about 2,000 cells/voxel at 7 T, comparable with gadolinium-labeled cells. Comparison of the detection sensitivity of cells labeled with (19)F, iron oxide and gadolinium over typical tissue background showed that unambiguous detection of the (19)F-labeled cells was simpler than with the contrast agents. The effect of the (19)F agent on cell function was minimal in the context of cell-based vaccines. From these data, we calculate that detection of 30,000 cells in vivo at 3 T with a reasonable signal to noise ratio for (19)F images would require less than 30 min with a conventional fast spin echo sequence, given a coil similar to the one used in this study. This is well within acceptable limits for clinical studies, and thus, we conclude that (19)F MRI for quantitative cell tracking in a clinical setting has great potential.

A novel (19)F agent for detection and quantification of human dendritic cells using magnetic resonance imaging.

We have demonstrated previously that BM-derived DCs can prevent diabetes development and halt progression of insulitis in NOD mice, the mouse model of type 1 diabetes. The DC population that was most effective in this therapy had a mature phenotype, expressed high levels of costimulatory molecules, and secreted low levels of IL-12p70. The protective DC therapy induced Treg and Th2 cells in vitro and in vivo. Microarray analysis of therapeutic and nontherapeutic DC populations revealed differences in the expression of OX40L, CD200, Ym-1, CCL2, and CCL5, which could play important roles in the observed DC-mediated therapy. The unique pattern of costimulatory molecules and chemokines expressed by the therapeutic DCs was confirmed by flow cytometry and ELISA. Using a novel cell-labeling and (19)F NMR, we observed that the chemokines secreted by the therapeutic DCs altered the migration of diabetogenic Th1 cells in vivo and attracted Th2 cells. These results suggest that the therapeutic function of DCs is mediated by a combination of costimulatory and chemokine properties that results in the attraction of diabetogenic Th1 and the induction of Th2 and/or Treg differentiation.

Gene expression analysis of dendritic cells that prevent diabetes in NOD mice: analysis of chemokines and costimulatory molecules.

Current biomarkers are unable to adequately predict vaccine-induced immune protection in humans with infectious disease or cancer. However, timely and adequate assessment of antigen-specific immune responses is critical for successful vaccine development. Therefore, we have developed a method for the direct assessment of immune responses in vivo in a clinical setting. Melanoma patients with lymph node (LN) metastases received dendritic cell (DC) vaccine therapy, injected intranodally, followed by [(18)F]-labeled 3'-fluoro-3'-deoxy-thymidine ([(18)F]FLT) PET at varying time points after vaccination. Control LNs received saline or DCs without antigen. De novo immune responses were readily visualized in treated LNs early after the prime vaccination, and these signals persisted for up to 3 wk. This selective [(18)F]FLT uptake was markedly absent in control LNs, although tracer uptake in treated LNs increased profoundly with as little as 4.5 × 10(5) DCs. Immunohistochemical staining confirmed injected DC dispersion to T-cell areas and resultant activation of CD4(+) and CD8(+) T cells. The level of LN tracer uptake significantly correlates to the level of circulating antigen-specific IgG antibodies and antigen-specific proliferation of T cells in peripheral blood. Furthermore, this correlation was not observed with [(18)F]-labeled fluoro-2-deoxy-2-D-glucose. Therefore, [(18)F]FLT PET offers a sensitive tool to study the kinetics, localization, and involvement of lymphocyte subsets in response to vaccination. This technique allows for early discrimination of responding from nonresponding patients in anti-cancer vaccination and aid physicians in individualized decisionmaking.

Early identification of antigen-specific immune responses in vivo by [18F]-labeled 3'-fluoro-3'-deoxy-thymidine ([18F]FLT) PET imaging.

In order for cellular therapeutics to succeed, comprehensive monitoring of the transplanted cells in vivo is required i.e., their localization, functionality and numbers in a longitudinal manner. Recently, dendritic cell based vaccines have been monitored by their effect on lymphocyte activation using [(18)F]FLT PET in cancer patients.

Insight into the dynamics, localization and magnitude of antigen-specific immune responses by [(18)F]FLT PET imaging.

Noninvasive in vivo cell tracking is crucial to fully understand the function of mobile and/or transplanted cells, particularly immune cells and cellular therapeutics. (19)F MRI for cell tracking has several advantages; chief among them are its noninvasive nature which allows longitudinal data acquisition, use of a stable, non-radioactive isotope permitting long-term tracking, the absence of confounding endogenous signal, and the ability to quantify cell numbers from image data. However, generation of sufficient signal i.e. (19)F cell loading is a key challenge, particularly with non-phagocytic cells such as lymphocytes and stem cells. A range of (19)F cell labels have been developed, including emulsions, particles, polymers, and agents for clinical use. Various animal and primary human cells, such as dendritic cells, lymphocytes and phagocytes have been successfully labeled and studied in models of autoimmune disease, inflammation and transplant rejection. Primary human cells, particularly dendritic cells as used in vaccine therapy have been tested for imminent clinical application. Here, we summarize current cell loading strategies and sensitivity of in vivo cell imaging with (19)F MRI, and discuss the processing of image data for accurate quantification of cell numbers. This novel technology is uniquely applicable to the longitudinal and quantitative tracking of cells in vivo.

Labeling cells for in vivo tracking using (19)F MRI.

&#x514D;&#x75AB;&#x6CBB;&#x7597;&#x7684;&#x76EE;&#x7684;&#x662F;&#x91CD;&#x65B0;&#x548C;&#x632F;&#x5174;&#x4E2D;&#x5BF9;&#x6297;&#x764C;&#x75C7;&#x7684;&#x514D;&#x75AB;&#x7CFB;&#x7EDF;&#x3002;&#x8FC7;&#x53BB;&#x51E0;&#x5341;&#x5E74;&#x7684;&#x7814;&#x7A76;&#x8868;&#x660E;&#x4EBA;&#x7C7B;&#x764C;&#x75C7;&#x4E0E;&#x514D;&#x75AB;&#x7CFB;&#x7EDF;&#x4E4B;&#x95F4;&#x7684;&#x5173;&#x7CFB;&#x662F;&#x590D;&#x6742;&#x3001; &#x9AD8;&#x5EA6;&#x52A8;&#x6001;&#x548C;&#x53D8;&#x91CF;&#x4E4B;&#x95F4;&#x7684;&#x4E2A;&#x4EBA;&#x3002;&#x8003;&#x8651;&#x590D;&#x6742;&#x6027;&#x3001; &#x4F5C;&#x51FA;&#x7684;&#x5DE8;&#x5927;&#x52AA;&#x529B;&#x548C;&#x4F18;&#x5316;&#x7684;&#x514D;&#x75AB;&#x6CBB;&#x7597;&#x65B9;&#x6CD5;&#x6240;&#x6D89;&#x7684;&#x8D39;&#x7528;&#xFF0C;&#x4E34;&#x5E8A;&#x4E0A;&#x9002;&#x7528;&#x7684;&#x5DE5;&#x5177;&#xFF0C;&#x4EE5;&#x76D1;&#x6D4B;&#x6CBB;&#x7597;&#x8BF1;&#x5BFC;&#x514D;&#x75AB;&#x53CD;&#x5E94;&#x4F53;&#x5185;&#x6700;&#x88AB;&#x5FC5;&#x8981;&#x3002;&#x7136;&#x800C;&#xFF0C;&#x8FD9;&#x7C7B;&#x5DE5;&#x5177;&#x7684;&#x53D1;&#x5C55;&#x88AB;&#x590D;&#x6742;&#x7684;&#x53D1;&#x5C55;&#x4E2D;&#x56FD;&#x5BB6;&#x7684;&#x514D;&#x75AB;&#x53CD;&#x5E94;&#x4F8B;&#x5982;&#x5305;&#x62EC;&#x8F66;&#x53A2;&#xFF0C;&#x8EAB;&#x4F53;&#x51E0;&#x4E2A;&#x4E8B;&#x5B9E;&#x3001; &#x5916;&#x5468;&#x7EC4;&#x7EC7;&#x3001; &#x6DCB;&#x5DF4;&#x7ED3;&#x3001; &#x6DCB;&#x5DF4;&#x7BA1;&#x548C;&#x8840;&#x7BA1;&#x7CFB;&#x7EDF;&#xFF0C;&#x4EE5;&#x53CA;&#x80BF;&#x7624;&#x7F51;&#x7AD9;&#x672C;&#x8EAB;&#x3002;&#x6B64;&#x5916;&#xFF0C;&#x5305;&#x62EC;&#x514D;&#x75AB;&#x7CFB;&#x7EDF;&#x7684;&#x7EC6;&#x80DE;&#x4E0D;&#x662F;&#x9759;&#x6001;&#x7684;&#x4F46;&#x4E0D;&#x65AD;&#x5FAA;&#x73AF;&#x7684;&#x8840;&#x7BA1;&#x548C;&#x6DCB;&#x5DF4;&#x7CFB;&#x7EDF;&#x3002;&#x5206;&#x5B50;&#x5F71;&#x50CF;&#x6280;&#x672F;&#x88AB;&#x8BA4;&#x4E3A;&#x662F;&#x6700;&#x559C;&#x6B22;&#x7684;&#x5019;&#x9009;&#x4EBA;&#xFF0C;&#x4EE5;&#x5C65;&#x884C;&#x8FD9;&#x4E00;&#x4EFB;&#x52A1;&#x3002;&#x6210;&#x50CF;&#x6280;&#x672F;&#x548C;&#x65B9;&#x5F0F;&#x53D6;&#x5F97;&#x8FDB;&#x5C55;&#x63D0;&#x4F9B;&#x4E86;&#x4E00;&#x4E2A;&#x591A;&#x529F;&#x80FD;&#x5DE5;&#x5177;&#x7BB1;&#x6765;&#x89E3;&#x51B3;&#x8FD9;&#x4E9B;&#x95EE;&#x9898;&#x3002;&#x8FD9;&#x4E00;&#x5BA1;&#x67E5;&#x4FA7;&#x91CD;&#x4E8E;&#x6CBB;&#x7597;&#x8BF1;&#x5BFC;&#x6297;&#x80BF;&#x7624;&#x514D;&#x75AB;&#x5E94;&#x7B54;&#x4F53;&#x5185;&#x68C0;&#x6D4B;&#x548C;&#x5168;&#x9762;&#x6982;&#x8FF0;&#x4E34;&#x5E8A;&#x4E0A;&#x53EF;&#x7528;&#x7684;&#x6210;&#x50CF;&#x6280;&#x672F;&#xFF0C;&#x4EE5;&#x53CA;&#x89C2;&#x70B9;&#x5BF9;&#x4ECA;&#x540E;&#x7684;&#x4E8B;&#x6001;&#x53D1;&#x5C55;&#x3002;&#x5728;&#x8BA8;&#x8BBA;&#x4E2D;&#xFF0C;&#x6211;&#x4EEC;&#x5C06;&#x4FA7;&#x91CD;&#x5177;&#x4F53;&#x6D89;&#x53CA;&#x5230;&#x514D;&#x75AB;&#x7CFB;&#x7EDF;&#x7684;&#x6210;&#x50CF;&#x7684;&#x95EE;&#x9898;&#x548C;&#x6211;&#x4EEC;&#x5C06;&#x8BA8;&#x8BBA;&#x7684;&#x957F;&#x5904;&#x548C;&#x5F53;&#x524D;&#x4E34;&#x5E8A;&#x6210;&#x50CF;&#x6280;&#x672F;&#x7684;&#x5C40;&#x9650;&#x6027;&#x3002;&#x6700;&#x540E;&#x4E00;&#x8282;&#x63D0;&#x4F9B;&#x4E86;&#x672A;&#x6765;&#x7684;&#x65B9;&#x5411;&#xFF0C;&#x6211;&#x4EEC;&#x8BBE;&#x60F3;&#x8981;&#x8FDB;&#x4E00;&#x6B65;&#x53D1;&#x5C55;&#x7684;&#x5173;&#x952E;&#x3002;

&#x8BF1;&#x5BFC;&#x6CBB;&#x7597;&#x4EBA;&#x7C7B;&#x6297;&#x764C;&#x514D;&#x75AB;&#x5E94;&#x7B54;&#x7684;&#x6D3B;&#x4F53;&#x6210;&#x50CF;&#x3002;

Immunotherapy aims to re-engage and revitalize the immune system in the fight against cancer. Research over the past decades has shown that the relationship between the immune system and human cancer is complex, highly dynamic, and variable between individuals. Considering the complexity, enormous effort and costs involved in optimizing immunotherapeutic approaches, clinically applicable tools to monitor therapy-induced immune responses in vivo are most warranted. However, the development of such tools is complicated by the fact that a developing immune response encompasses several body compartments, e.g., peripheral tissues, lymph nodes, lymphatic and vascular systems, as well as the tumor site itself. Moreover, the cells that comprise the immune system are not static but constantly circulate through the vascular and lymphatic system. Molecular imaging is considered the favorite candidate to fulfill this task. The progress in imaging technologies and modalities has provided a versatile toolbox to address these issues. This review focuses on the detection of therapy-induced anticancer immune responses in vivo and provides a comprehensive overview of clinically available imaging techniques as well as perspectives on future developments. In the discussion, we will focus on issues that specifically relate to imaging of the immune system and we will discuss the strengths and limitations of the current clinical imaging techniques. The last section provides future directions that we envision to be crucial for further development.

In vivo imaging of therapy-induced anti-cancer immune responses in humans.

Immunoth&eacute;rapie vise &agrave; r&eacute;engager et revitaliser le syst&egrave;me immunitaire dans la lutte contre le cancer. Recherches durant les derni&egrave;res d&eacute;cennies ont montr&eacute; que la relation entre le syst&egrave;me immunitaire et le cancer humain est complexe, tr&egrave;s dynamique et variable entre les individus. Compte tenu de la complexit&eacute;, des efforts consid&eacute;rables et co&ucirc;ts li&eacute;s &agrave; l&#39;optimisation des approches immunoth&eacute;rapeutiques, outils cliniquement il y a lieu de surveiller les r&eacute;ponses d&#39;immunitaire induite par le traitement in vivo sont plus garantis. Cependant, le d&eacute;veloppement de ces outils est compliqu&eacute; par le fait qu&#39;une r&eacute;ponse immunitaire en d&eacute;veloppement englobe plusieurs compartiments de l&#39;organisme, par exemple, les tissus p&eacute;riph&eacute;riques, ganglions lymphatiques, des syst&egrave;mes vasculaires et lymphatiques, ainsi que le site de la tumeur elle-m&ecirc;me. En outre, les cellules qui composent le syst&egrave;me immunitaire ne sont pas statiques mais circulent constamment dans le syst&egrave;me vasculaire et lymphatique. Imagerie mol&eacute;culaire est consid&eacute;r&eacute; comme le candidat favori pour remplir cette mission. L&#39;&eacute;tat d&#39;avancement des technologies et des modalit&eacute;s d&#39;imagerie a fourni une bo&icirc;te &agrave; outils polyvalent pour r&eacute;soudre ces probl&egrave;mes. Cette revue se concentre sur la d&eacute;tection des induite par la th&eacute;rapie anticanc&eacute;reuses r&eacute;ponses immunitaires in vivo et fournit une vue d&#39;ensemble des techniques d&#39;imagerie disponibles sur le plan clinique, mais aussi des perspectives sur les d&eacute;veloppements futurs. Dans la discussion, nous nous concentrerons sur les questions qui concernent sp&eacute;cifiquement l&#39;imagerie du syst&egrave;me immunitaire et nous allons discuter les points forts et limites des techniques d&#39;imagerie cliniques actuelles. La derni&egrave;re section fournit les orientations futures que nous envisageons pour &ecirc;tre crucial pour le d&eacute;veloppement ult&eacute;rieur.

Imagerie in vivo d&#39;induite par le traitement des r&eacute;ponses immunitaires contre le cancer chez les humains.

Immuntherapie soll selbstm&ouml;rderischen und beleben das Immunsystem im Kampf gegen den Krebs. Forschung in den letzten Jahrzehnten hat gezeigt, dass die Beziehung zwischen dem Immunsystem und dem menschlichen Krebs Komplex, sehr dynamisch und variabel zwischen Individuen. In Anbetracht der Komplexit&auml;t, enormen Aufwand und Kosten optimieren immuntherapeutischen Ans&auml;tze vorliegen, werden die meisten klinisch anwendbare Werkzeuge zur &Uuml;berwachung der Therapie-induzierte Immunantworten in-vivo. Jedoch ist die Entwicklung solcher Werkzeuge erschwert durch die Tatsache, dass eine entwickelnde Immune Antwort mehrere K&ouml;rper-Abteile, z.B. umfasst, peripheren Geweben, Lymphknoten, Lymph- und vaskul&auml;re Systeme, als auch die Tumor-Website selbst. Dar&uuml;ber hinaus die Zellen, die das Immunsystem bilden sind nicht statisch, sondern st&auml;ndig durch die Gef&auml;&szlig;- und lymphatische System zirkulieren. Molekulare Bildgebung gilt als den bevorzugten Kandidaten f&uuml;r diese Aufgabe zu erf&uuml;llen. Die Fortschritte bei der imaging-Technologien und Modalit&auml;ten lieferte ein vielseitiges Instrumentarium um diese Probleme anzugehen. Diesen Bericht konzentriert sich auf die Erkennung von Therapie-induzierte krebsbek&auml;mpfende Immunantworten in-vivo und bietet einen umfassenden &Uuml;berblick &uuml;ber klinisch verf&uuml;gbaren bildgebenden Verfahren sowie Perspektiven auf zuk&uuml;nftige Entwicklungen. In der Diskussion wir konzentrieren uns auf Fragen, die speziell auf die Bildgebung des Immunsystems beziehen, und wir werden diskutieren, die St&auml;rken und Schw&auml;chen der aktuellen klinischen bildgebender Verfahren. Das letzte Kapitel bietet Zukunftsperspektiven, die wir vorstellen, um f&uuml;r die weitere Entwicklung von entscheidender Bedeutung sein.

In Vivo Bildgebung und Therapie-induzierte krebshemmenden Immunantworten beim Menschen.

Immunoterapia mira a coinvolgere nuovamente e rivitalizzare il sistema immunitario nella lotta contro il cancro. Ricerca negli ultimi decenni ha dimostrato che il rapporto tra il sistema immunitario e il cancro umano &egrave; complesso, altamente dinamico e variabile tra gli individui. Considerando la complessit&agrave;, enormi sforzi e costi di ottimizzazione approcci immunotherapeutic, clinicamente applicabili strumenti per monitorare la terapia-indotta immune responses in vivo sono pi&ugrave; garantiti. Tuttavia, lo sviluppo di tali strumenti &egrave; complicato dal fatto che una risposta immunitaria in via di sviluppo comprende diversi compartimenti del corpo, ad esempio, tessuti periferici, i linfonodi, sistemi linfatici e vascolari, come pure il sito del tumore stesso. Inoltre, le cellule che compongono il sistema immunitario non sono statiche ma costantemente circolano attraverso il sistema vascolare e linfatico. Imaging molecolare &egrave; considerato il candidato favorito per adempiere questo compito. I progressi nelle tecnologie di imaging e modalit&agrave; ha fornito una serie di strumenti versatili per risolvere questi problemi. Questa recensione si concentra sull&#39;individuazione di indotta da terapia antitumorale risposte immunitarie in vivo e fornisce una panoramica completa delle tecniche di imaging clinicamente disponibile come pure prospettive sugli sviluppi futuri. Nella discussione, ci concentreremo su questioni che si riferiscono specificamente all&#39;imaging del sistema immunitario e discuteremo i punti di forza e limiti delle tecniche di imaging cliniche attuale. L&#39;ultima sezione fornisce indicazioni future che immaginiamo per essere cruciale per un ulteriore sviluppo.

Imaging in vivo di indotta da terapia anti-cancro immune responses in esseri umani.

&#x514D;&#x75AB;&#x7642;&#x6CD5;&#x5F93;&#x4E8B;&#x3057;&#x3001;&#x304C;&#x3093;&#x3068;&#x306E;&#x6226;&#x3044;&#x3067;&#x306E;&#x514D;&#x75AB;&#x30B7;&#x30B9;&#x30C6;&#x30E0;&#x306E;&#x6D3B;&#x6027;&#x5316;&#x3092;&#x76EE;&#x6307;&#x3057;&#x3066;&#x3044;&#x307E;&#x3059;&#x3002;&#x904E;&#x53BB;&#x5341;&#x5E74;&#x306B;&#x308F;&#x305F;&#x3063;&#x3066;&#x7814;&#x7A76;&#x306F;&#x3001;&#x514D;&#x75AB;&#x30B7;&#x30B9;&#x30C6;&#x30E0;&#x3068;&#x4EBA;&#x9593;&#x306E;&#x304C;&#x3093;&#x3068;&#x306E;&#x95A2;&#x4FC2;&#x304C;&#x8907;&#x96D1;&#x306A;&#x975E;&#x5E38;&#x306B;&#x30C0;&#x30A4;&#x30CA;&#x30DF;&#x30C3;&#x30AF;&#x306A;&#x500B;&#x4EBA;&#x9593;&#x306E;&#x5909;&#x6570;&#x3067;&#x3042;&#x308B;&#x3053;&#x3068;&#x3092;&#x793A;&#x3057;&#x3066;&#x3044;&#x307E;&#x3059;&#x3002;&#x7642;&#x6CD5;&#x306B;&#x3088;&#x308B;&#x514D;&#x75AB;&#x5FDC;&#x7B54;&#x306E; in vivo &#x3092;&#x76E3;&#x8996;&#x3059;&#x308B;&#x305F;&#x3081;&#x306E;&#x81E8;&#x5E8A;&#x5FDC;&#x7528;&#x53EF;&#x80FD;&#x306A;&#x30C4;&#x30FC;&#x30EB;&#x306F;&#x3001;&#x8907;&#x96D1;&#x3055;&#x306B;&#x591A;&#x5927;&#x306A;&#x52B4;&#x529B;&#x3068;&#x30B3;&#x30B9;&#x30C8;&#x6700;&#x9069;&#x5316;&#x306E;&#x514D;&#x75AB;&#x7642;&#x6CD5;&#x30A2;&#x30D7;&#x30ED;&#x30FC;&#x30C1;&#x306E;&#x95A2;&#x4E0E;&#x3092;&#x8003;&#x3048;&#x308B;&#x6700;&#x3082;&#x4FDD;&#x8A3C;&#x3055;&#x308C;&#x307E;&#x3059;&#x3002;&#x305F;&#x3060;&#x3057;&#x3001;&#x3053;&#x306E;&#x3088;&#x3046;&#x306A;&#x30C4;&#x30FC;&#x30EB;&#x306E;&#x958B;&#x767A;&#x306F;&#x514D;&#x75AB;&#x5FDC;&#x7B54;&#x3092;&#x958B;&#x767A;&#x3057;&#x3066;&#x4F53;&#x306E;&#x8907;&#x6570;&#x306E;&#x4ED5;&#x5207;&#x308A;&#x3092;&#x4F8B;&#x3048;&#x3070;&#x5305;&#x542B;&#x3059;&#x308B;&#x3053;&#x3068;&#x4E8B;&#x5B9F;&#x3001;&#x672B;&#x68A2;&#x7D44;&#x7E54;&#x3001;&#x30EA;&#x30F3;&#x30D1;&#x7BC0;&#x3001;&#x30EA;&#x30F3;&#x30D1;&#x304A;&#x3088;&#x3073;&#x8840;&#x7BA1;&#x30B7;&#x30B9;&#x30C6;&#x30E0;&#x3068;&#x540C;&#x69D8;&#x306B;&#x3088;&#x308B;&#x816B;&#x760D;&#x90E8;&#x4F4D;&#x81EA;&#x4F53;&#x8907;&#x96D1;&#x3067;&#x3059;&#x3002;&#x307E;&#x305F;&#x3001;&#x514D;&#x75AB;&#x30B7;&#x30B9;&#x30C6;&#x30E0;&#x3092;&#x69CB;&#x6210;&#x3059;&#x308B;&#x7D30;&#x80DE;&#x306F;&#x9759;&#x7684;&#x3067;&#x306F;&#x3042;&#x308A;&#x307E;&#x305B;&#x3093;&#x304C;&#x3001;&#x5E38;&#x306B;&#x8840;&#x7BA1;&#x3068;&#x30EA;&#x30F3;&#x30D1;&#x7CFB;&#x3092;&#x901A;&#x3057;&#x3066;&#x5FAA;&#x74B0;&#x3057;&#x307E;&#x3059;&#x3002;&#x5206;&#x5B50;&#x30A4;&#x30E1;&#x30FC;&#x30B8;&#x30F3;&#x30B0;&#x306F;&#x3001;&#x3053;&#x306E;&#x30BF;&#x30B9;&#x30AF;&#x3092;&#x9054;&#x6210;&#x3059;&#x308B;&#x305F;&#x3081;&#x306B;&#x304A;&#x6C17;&#x306B;&#x5165;&#x308A;&#x306E;&#x5019;&#x88DC;&#x3068;&#x898B;&#x306A;&#x3055;&#x308C;&#x307E;&#x3059;&#x3002;&#x30A4;&#x30E1;&#x30FC;&#x30B8;&#x30F3;&#x30B0;&#x6280;&#x8853;&#x3068;&#x30E2;&#x30C0;&#x30EA;&#x30C6;&#x30A3;&#x306E;&#x4E2D;&#x306F;&#x3001;&#x3053;&#x308C;&#x3089;&#x306E;&#x554F;&#x984C;&#x306B;&#x5BFE;&#x51E6;&#x3059;&#x308B;&#x591A;&#x76EE;&#x7684;&#x306A;&#x30C4;&#x30FC;&#x30EB;&#x30DC;&#x30C3;&#x30AF;&#x30B9;&#x3092;&#x63D0;&#x4F9B;&#x3057;&#x3066;&#x3044;&#x307E;&#x3059;&#x3002;&#x3053;&#x306E;&#x30EC;&#x30D3;&#x30E5;&#x30FC;&#x306F;&#x7642;&#x6CD5;&#x306B;&#x3088;&#x308B;&#x6297;&#x304C;&#x3093;&#x514D;&#x75AB;&#x5FDC;&#x7B54; in vivo &#x691C;&#x51FA;&#x306B;&#x7126;&#x70B9;&#x3092;&#x5F53;&#x3066;&#x3001;&#x5C06;&#x6765;&#x306E;&#x767A;&#x5C55;&#x306B;&#x898B;&#x65B9;&#x3068;&#x540C;&#x69D8;&#x306B;&#x81E8;&#x5E8A;&#x7684;&#x306B;&#x5229;&#x7528;&#x53EF;&#x80FD;&#x306A;&#x30A4;&#x30E1;&#x30FC;&#x30B8;&#x30F3;&#x30B0;&#x6280;&#x8853;&#x306E;&#x5305;&#x62EC;&#x7684;&#x306A;&#x6982;&#x8981;&#x3057;&#x307E;&#x3059;&#x3002;&#x30C7;&#x30A3;&#x30B9;&#x30AB;&#x30C3;&#x30B7;&#x30E7;&#x30F3;&#x306F;&#x3001;&#x79C1;&#x305F;&#x3061;&#x306E;&#x514D;&#x75AB;&#x30B7;&#x30B9;&#x30C6;&#x30E0;&#x306E;&#x30A4;&#x30E1;&#x30FC;&#x30B8;&#x30F3;&#x30B0;&#x306B;&#x5177;&#x4F53;&#x7684;&#x306B;&#x95A2;&#x9023;&#x3059;&#x308B;&#x554F;&#x984C;&#x306B;&#x7126;&#x70B9;&#x3092;&#x5F53;&#x3066;&#x308B;&#x3001;&#x6211;&#x3005; &#x306E;&#x5F37;&#x307F;&#x3068;&#x73FE;&#x5728;&#x306E;&#x81E8;&#x5E8A;&#x753B;&#x50CF;&#x51E6;&#x7406;&#x6280;&#x8853;&#x306E;&#x9650;&#x754C;&#x306B;&#x3064;&#x3044;&#x3066;&#x8AAC;&#x660E;&#x3057;&#x307E;&#x3059;&#x3002;&#x6700;&#x5F8C;&#x306B;&#x3001;&#x79C1;&#x9054;&#x306F;&#x958B;&#x767A;&#x306E;&#x305F;&#x3081;&#x306B;&#x3055;&#x3089;&#x306B;&#x91CD;&#x8981;&#x3067;&#x3042;&#x308B;&#x3053;&#x3068;&#x3092;&#x60F3;&#x50CF;&#x5C06;&#x6765;&#x306E;&#x65B9;&#x5411;&#x6027;&#x306B;&#x3064;&#x3044;&#x3066;&#x8AAC;&#x660E;&#x3057;&#x307E;&#x3059;&#x3002;

In vivo &#x30A4;&#x30E1;&#x30FC;&#x30B8;&#x30F3;&#x30B0;&#x7642;&#x6CD5;&#x8A98;&#x767A;&#x6297;&#x304C;&#x3093;&#x514D;&#x75AB;&#x4EBA;&#x9593;&#x306E;&#x3002;

Immunotherapy &#xAD50;&#xC804; &#xD558; &#xACE0; &#xC554;&#xACFC;&#xC758; &#xC2F8;&#xC6C0;&#xC5D0;&#xC11C; &#xBA74;&#xC5ED; &#xC2DC;&#xC2A4;&#xD15C;&#xC744; &#xD65C;&#xC131;&#xD654; &#xD558;&#xB294; &#xAC83;&#xC744; &#xBAA9;&#xD45C;&#xB85C;. &#xC9C0;&#xB09C; &#xC218; &#xC2ED;&#xB144; &#xB3D9;&#xC548; &#xC5F0;&#xAD6C;&#xB294; &#xBA74;&#xC5ED; &#xCCB4;&#xACC4;&#xC640; &#xC778;&#xAC04;&#xC758; &#xC554; &#xC0AC;&#xC774; &#xAD00;&#xACC4;&#xB294; &#xB9E4;&#xC6B0; &#xB3D9;&#xC801; &#xBCF5;&#xC7A1; &#xD558; &#xACE0; &#xAC1C;&#xC778; &#xC0AC;&#xC774;&#xC758; &#xAC00;&#xBCC0; &#xBCF4;&#xC774;&#xACE0; &#xC788;&#xB2E4;. &#xBCF5;&#xC7A1;&#xC131;, &#xAC70; &#xB300; &#xD55C; &#xB178;&#xB825; &#xBC0F; &#xCD5C;&#xC801;&#xD654; immunotherapeutic &#xC811;&#xADFC;&#xC5D0; &#xAD00;&#xB828; &#xB41C; &#xBE44;&#xC6A9;&#xC744; &#xACE0;&#xB824; &#xD558;&#xBA74; &#xCE58;&#xB8CC; &#xC720;&#xB3C4; &#xB41C; &#xBA74;&#xC5ED; &#xBC18;&#xC751; &#xC0DD;&#xCCB4; &#xC870;&#xAC74; &#xBAA8;&#xB2C8;&#xD130;&#xB9C1; &#xD558;&#xB294; &#xB3C4;&#xAD6C;&#xB97C; &#xC784;&#xC0C1;&#xC801;&#xC73C;&#xB85C; &#xC801;&#xC6A9; &#xAC00;&#xB2A5;&#xD55C; &#xAC00;&#xC7A5; &#xBCF4;&#xC99D; &#xD569;&#xB2C8;&#xB2E4;. &#xADF8;&#xB7EC;&#xB098;, &#xC774;&#xB7EC;&#xD55C; &#xB3C4;&#xAD6C; &#xAC1C;&#xBC1C; &#xAC1C;&#xBC1C; &#xBA74;&#xC5ED; &#xBC18;&#xC751; &#xC608;&#xB97C; &#xB4E4;&#xC5B4; &#xBA87; &#xAC00;&#xC9C0; &#xC2E0;&#xCCB4; &#xAD6C;&#xD68D;&#xB97C; &#xD3EC;&#xD568; &#xD558;&#xB294; &#xC0AC;&#xC2E4;, &#xC8FC;&#xBCC0; &#xC870;&#xC9C1;, &#xB9BC;&#xD504;&#xC808;, &#xB9BC;&#xD504; &#xBC0F; &#xD608;&#xAD00; &#xC2DC;&#xC2A4;&#xD15C; &#xBFD0;&#xB9CC; &#xC544;&#xB2C8;&#xB77C; &#xC885;&#xC591; &#xC0AC;&#xC774;&#xD2B8; &#xC790;&#xCCB4;&#xC5D0; &#xC758;&#xD574; &#xBCF5;&#xC7A1; &#xD569;&#xB2C8;&#xB2E4;. &#xB610;&#xD55C;, &#xBA74;&#xC5ED; &#xCCB4;&#xACC4;&#xB97C; &#xAD6C;&#xC131; &#xD558;&#xB294; &#xC140;&#xC740; &#xC815;&#xC801; &#xD558;&#xC9C0;&#xB9CC; &#xC9C0;&#xC18D;&#xC801;&#xC73C;&#xB85C; &#xD608;&#xAD00;&#xACFC; &#xB9BC;&#xD504; &#xC2DC;&#xC2A4;&#xD15C;&#xC744; &#xD1B5;&#xD574; &#xC21C;&#xD658;. &#xBD84;&#xC790; &#xD654;&#xC0C1; &#xC9C4; &#xCC30;&#xC5D0;&#xB294;&#xC774; &#xC791;&#xC5C5;&#xC744; &#xC218;&#xD589; &#xD558;&#xAE30; &#xC704;&#xD574; &#xC88B;&#xC544;&#xD558;&#xB294; &#xD6C4;&#xBCF4;&#xB85C; &#xAC04;&#xC8FC; &#xB429;&#xB2C8;&#xB2E4;. &#xC774;&#xBBF8;&#xC9D5; &#xAE30;&#xC220;&#xACFC; modalities&#xC5D0; &#xC9C4;&#xD589; &#xC774;&#xB7EC;&#xD55C; &#xBB38;&#xC81C;&#xB97C; &#xD574;&#xACB0; &#xD558;&#xAE30; &#xC704;&#xD574; &#xB2E4;&#xC591; &#xD55C; &#xB3C4;&#xAD6C;&#xB97C; &#xC81C;&#xACF5; &#xD558;&#xACE0;&#xC788;&#xB2E4;. &#xC774; &#xAC80;&#xD1A0; &#xCE58;&#xB8CC; &#xC720;&#xB3C4; &#xD56D; &#xC554; &#xBA74;&#xC5ED; &#xBC18;&#xC751; &#xC0DD;&#xCCB4; &#xC870;&#xAC74; &#xAC80;&#xC0C9;&#xC5D0; &#xCD08;&#xC810;&#xC744; &#xB9DE;&#xCD94;&#xACE0; &#xBC0F; &#xBBF8;&#xB798;&#xC758; &#xBC1C;&#xC804;&#xC5D0; &#xC784;&#xC0C1;&#xC801;&#xC73C;&#xB85C; &#xC0AC;&#xC6A9;&#xD560; &#xC218; &#xC788;&#xB294; &#xC774;&#xBBF8;&#xC9D5; &#xAE30;&#xBC95; &#xAD00;&#xC810;&#xC758; &#xD3EC;&#xAD04;&#xC801;&#xC778; &#xAC1C;&#xC694;&#xB97C; &#xC81C;&#xACF5; &#xD569;&#xB2C8;&#xB2E4;. &#xD1A0;&#xB860;&#xC5D0;&#xC11C; &#xC6B0;&#xB9AC;&#xB294; &#xD2B9;&#xD788; &#xBA74;&#xC5ED; &#xC2DC;&#xC2A4;&#xD15C;&#xC758; &#xC774;&#xBBF8;&#xC9D5;&#xC5D0; &#xAD00;&#xB828; &#xB41C; &#xBB38;&#xC81C;&#xC5D0; &#xCD08;&#xC810; &#xBC0F; &#xC7A5;&#xC810;&#xACFC; &#xD604;&#xC7AC; &#xC784;&#xC0C1; &#xC774;&#xBBF8;&#xC9D5; &#xAE30;&#xC220;&#xC758; &#xD55C;&#xACC4;&#xB97C; &#xC0B4;&#xD3B4;&#xBCFC; &#xAC83;. &#xB9C8;&#xC9C0;&#xB9C9; &#xC139;&#xC158;&#xC5D0;&#xC11C; &#xCD94;&#xAC00; &#xAC1C;&#xBC1C;&#xC744; &#xC704;&#xD55C; &#xC911;&#xC694; &#xD55C; &#xC6B0;&#xB9AC;&#xB97C; &#xAD6C;&#xC0C1; &#xD558;&#xB294; &#xBBF8;&#xB798;&#xC758; &#xBC29;&#xD5A5;&#xC744; &#xC81C;&#xACF5; &#xD569;&#xB2C8;&#xB2E4;.

&#xC778; &#xAC04;&#xC5D0; &#xC788;&#xB294; &#xD56D; &#xC554; &#xBA74;&#xC5ED; &#xBC18;&#xC751;&#xC744; &#xCE58;&#xB8CC; &#xC720;&#xB3C4;&#xC758; in vivo &#xC601;&#xC0C1;.

La inmunoterapia pretende entablar nuevos contactos y revitalizar el sistema inmunol&oacute;gico en la lucha contra el c&aacute;ncer. Investigaci&oacute;n en las &uacute;ltimas d&eacute;cadas ha demostrado que la relaci&oacute;n entre el sistema inmune y c&aacute;ncer en humanos es complejo, din&aacute;mico y variable entre individuos. Teniendo en cuenta la complejidad, un esfuerzo enorme y los costos involucrados en enfoques inmunoterap&eacute;uticos optimizando, cl&iacute;nicamente aplicables herramientas para monitorear la terapia-respuestas inmunitarias inducidas por in vivo m&aacute;s est&aacute;n garantizados. Sin embargo, el desarrollo de estas herramientas se complica por el hecho de que una respuesta inmune en desarrollo abarca varios compartimientos del cuerpo, por ejemplo, los tejidos perif&eacute;ricos, ganglios linf&aacute;ticos, sistemas linf&aacute;ticos y vasculares, as&iacute; como el sitio del tumor. Por otra parte, las c&eacute;lulas que componen el sistema inmunitario no son est&aacute;ticas, pero constantemente circulan a trav&eacute;s del sistema linf&aacute;tico y vascular. Imagen molecular es considerado el candidato favorito para cumplir con esta tarea. Los avances en tecnolog&iacute;as y modalidades ha proporcionado una caja de herramientas vers&aacute;til para tratar estos temas. Esta revisi&oacute;n se centra en la detecci&oacute;n de inducida por terapia antic&aacute;ncer inmunorespuestas in vivo y proporciona una visi&oacute;n completa de las t&eacute;cnicas de imagen cl&iacute;nicamente disponibles as&iacute; como perspectivas sobre la evoluci&oacute;n futura. En la discusi&oacute;n, nos centraremos en las cuestiones que se refieren espec&iacute;ficamente a la proyecci&oacute;n de imagen del sistema inmune y discutiremos las fortalezas y limitaciones de las t&eacute;cnicas de imagen cl&iacute;nicas actuales. La &uacute;ltima secci&oacute;n proporciona el futuro que queremos para ser crucial para el desarrollo.

Imagen en vivo de inducida por terapia antic&aacute;ncer inmunorespuestas en seres humanos.

Cellular therapy can be defined as the transplantation of living cells for the treatment of medical conditions. Three main objectives of cellular therapy are regeneration of damaged tissue, replacement of function by secretion of biologically active molecules, and redirection of aberrant processes. Given the complex nature of these approaches, in vivo tracking of the transplanted cells is critical to evaluate their potential benefit and to optimize treatment strategies. Recent advances are reviewed that enable in vivo cell tracking as an important adjunct to implement cellular therapy in clinical practice.

In vivo tracking techniques for cellular regeneration, replacement, and redirection.

Anticancer dendritic cell (DC) vaccines require the DCs to relocate to lymph nodes (LN) to trigger immune responses. However, these migration rates are typically very poor. Improving the targeting of ex vivo generated DCs to LNs might increase vaccine efficacy and reduce costs. We investigated DC migration in vivo in humans under different conditions.

Targeting of 111In-labeled dendritic cell human vaccines improved by reducing number of cells.

Dendritic cell (DC)-based vaccines require the cells to relocate to lymph nodes (LNs). Unfortunately, however, DC migration rates are typically very poor. We investigated strategies to increase the migration efficacy of DC-based vaccines. Surprisingly, a reduction in DC number, but not the conditioning of the injection site, improved LN targeting.

Article	Year
Magnetic resonance imaging of embryonic and fetal development in model systems. Methods in molecular medicine\| PubMed ID: 16506418	2006
Fluorine-19 MRI for visualization and quantification of cell migration in a diabetes model. Magnetic resonance in medicine : official journal of the Society of Magnetic Resonance in Medicine / Society of Magnetic Resonance in Medicine\| PubMed ID: 17899609	2007
Self-delivering nanoemulsions for dual fluorine-19 MRI and fluorescence detection. Journal of the American Chemical Society\| PubMed ID: 18266363	2008
In vivo cytometry of antigen-specific t cells using 19F MRI. Magnetic resonance in medicine : official journal of the Society of Magnetic Resonance in Medicine / Society of Magnetic Resonance in Medicine\| PubMed ID: 19585593	2009
(19)F MRI for quantitative in vivo cell tracking. Trends in biotechnology\| PubMed ID: 20427096	2010
Customizable, multi-functional fluorocarbon nanoparticles for quantitative in vivo imaging using 19F MRI and optical imaging. Biomaterials\| PubMed ID: 20566214	2010
A novel (19)F agent for detection and quantification of human dendritic cells using magnetic resonance imaging. International journal of cancer. Journal international du cancer\| PubMed ID: 20839261	2011
Gene expression analysis of dendritic cells that prevent diabetes in NOD mice: analysis of chemokines and costimulatory molecules. Journal of leukocyte biology\| PubMed ID: 21628331	2011
Early identification of antigen-specific immune responses in vivo by [18F]-labeled 3'-fluoro-3'-deoxy-thymidine ([18F]FLT) PET imaging. Proceedings of the National Academy of Sciences of the United States of America\| PubMed ID: 22025695	2011
Insight into the dynamics, localization and magnitude of antigen-specific immune responses by [(18)F]FLT PET imaging. Oncoimmunology\| PubMed ID: 22934268	2012
Labeling cells for in vivo tracking using (19)F MRI. Biomaterials\| PubMed ID: 22959182	2012
In vivo imaging of therapy-induced anti-cancer immune responses in humans. Cellular and molecular life sciences : CMLS\| PubMed ID: 23052208	2012
In vivo tracking techniques for cellular regeneration, replacement, and redirection. Journal of nuclear medicine : official publication, Society of Nuclear Medicine\| PubMed ID: 23143090	2012
Targeting of 111In-labeled dendritic cell human vaccines improved by reducing number of cells. Clinical cancer research : an official journal of the American Association for Cancer Research\| PubMed ID: 23382117	2013
Reducing cell number improves the homing of dendritic cells to lymph nodes upon intradermal vaccination. Oncoimmunology\| PubMed ID: 24073362	2013

Mangala Srinivas

Biography

JoVE Journal Publications

Other Publications