Cell Tracking Methods Research Articles

In Vivo Cell Tracking Using PET: Opportunities and Challenges for Clinical Translation in Oncology.

Simple SummaryTracking therapeutic cells with non-invasive imaging methods has the potential to provide important information on the efficacy of cell therapies. In oncology, for example, monitoring the spatial distribution of chimeric antigen receptor (CAR) T-cells or tumour-infiltrating lymphocytes (TILs) could be used to monitor the efficiency of cellular trafficking to target sites within a patient. This review covers different cell labelling approaches for the non-invasive detection of therapeutic cells using positron emission tomography (PET). The potential for the clinical translation of these approaches and first-in-human studies is examined, as well as the translational challenges involved and how imaging can help overcome some of these challenges.Cell therapy is a rapidly evolving field involving a wide spectrum of therapeutic cells for personalised medicine in cancer. In vivo imaging and tracking of cells can provide useful information for improving the accuracy, efficacy, and safety of cell therapies. This review focuses on radiopharmaceuticals for the non-invasive detection and tracking of therapeutic cells using positron emission tomography (PET). A range of approaches for imaging therapeutic cells is discussed: Direct ex vivo labelling of cells, in vivo indirect labelling of cells by utilising gene reporters, and detection of specific antigens expressed on the target cells using antibody-based radiopharmaceuticals (immuno-PET). This review examines the evaluation of PET imaging methods for therapeutic cell tracking in preclinical cancer models, their role in the translation into patients, first-in-human studies, as well as the translational challenges involved and how they can be overcome.

Tracking leukocytes in intravital time lapse images using 3D cell association learning network

Leukocytes are key cellular elements of the innate immune system in all vertebrates, which play a crucial role in defending organisms against invading pathogens. Tracking these highly migratory and amorphous cells in in vivo models such as zebrafish embryos is a challenging task in cellular immunology. As temporal and special analysis of these imaging datasets by a human operator is quite laborious, developing an automated cell tracking method is highly in demand. Despite the remarkable advances in cell detection, this field still lacks powerful algorithms to accurately associate the detected cell across time frames. The cell association challenge is mostly related to the amorphous nature of cells, and their complicated motion profile through their migratory paths. To tackle the cell association challenge, we proposed a novel deep-learning-based object linkage method. For this aim, we trained the 3D cell association learning network (3D-CALN) with enough manually labelled paired 3D images of single fluorescent zebrafish's neutrophils from two consecutive frames. Our experiment results prove that deep learning is significantly applicable in cell linkage and particularly for tracking highly mobile and amorphous leukocytes. A comparison of our tracking accuracy with other available tracking algorithms shows that our approach performs well in relation to addressing cell tracking problems.

A Novel Method for Effective Cell Segmentation and Tracking in Phase Contrast Microscopic Images.

Cell migration plays an important role in the identification of various diseases and physiological phenomena in living organisms, such as cancer metastasis, nerve development, immune function, wound healing, and embryo formulation and development. The study of cell migration with a real-time microscope generally takes several hours and involves analysis of the movement characteristics by tracking the positions of cells at each time interval in the images of the observed cells. Morphological analysis considers the shapes of the cells, and a phase contrast microscope is used to observe the shape clearly. Therefore, we developed a segmentation and tracking method to perform a kinetic analysis by considering the morphological transformation of cells. The main features of the algorithm are noise reduction using a block-matching 3D filtering method, k-means clustering to mitigate the halo signal that interferes with cell segmentation, and the detection of cell boundaries via active contours, which is an excellent way to detect boundaries. The reliability of the algorithm developed in this study was verified using a comparison with the manual tracking results. In addition, the segmentation results were compared to our method with unsupervised state-of-the-art methods to verify the proposed segmentation process. As a result of the study, the proposed method had a lower error of less than 40% compared to the conventional active contour method.

A novel imaging tool; fluorescent organosilica nanoparticles

The novel type of silica nanoparticles, organosilica nanoparticles, has been developed by Nakamura and colleagues. The nanoparticles can be synthesized in a few of simple reactions and have a good size and dispersibility. Organosilica nanoparticles have SH groups on their surface, which makes it possible to conjugate them with antibodies and other protein molecules. We synthesized organosilica nanoparticles with a diameter of 100 nm, containing fluorescein isothiocyanate (FITC) or Rhodamine B, which resulted in high fluorescence intensity. These nanoparticles were tested in various biochemical applications and in this study, we focused on their use in flow cytometry. We found that the application of these nanoparticles with antibodies could efficiently detect specific cells in a flow cytometer. We also discuss cell tracking methods using these nanoparticles.

Non-invasive synchronous monitoring of neutrophil migration using whole body near-infrared fluorescence-based imaging

Advances in fluorescence imaging coupled with the generation of near infrared probes have significantly improved the capabilities of non-invasive, real-time imaging in whole animals. In this study we were able to overcome a limitation of in vivo fluorescence imaging and have established a dual cell tracking method where two different cell types can be monitored according to the spectral signature of the cell labelling fluorophore. Using a mouse model of acute liver injury, we have characterised the in vivo migration patterns of wild type and transgenic neutrophils with impaired chemotaxis. Here, we were able to demonstrate that IVIS provides a sensitive multiplexing technology to differentiate two different cell populations based on the spectral signature of the cell labelling fluorophores. This spectral unmixing methodology has the potential to uncover multidimensional cellular interactions involved in many diseases such as fibrosis and cancer. In vivo spectral un-mixing provides a useful tool for monitoring multiple biological process in real-time in the same animal.

Tracking Longitudinal Population Dynamics of Single Neuronal Calcium Signal Using SCOUT

In vivo calcium imaging enables simultaneous recording of large neuronal ensembles engaged in complex operations. Many experiments require monitoring and identification of cell populations across multiple sessions. Population cell tracking across multiple sessions is complicated by non-rigid transformations induced by cell movement and imaging field shifts. We introduce SCOUT (Single-Cell SpatiOtemporal LongitUdinal Tracking), a fast, robust cell tracking method utilizing multiple cell-cell similarity metrics, probabilistic inference, and an adaptive clustering methodology to perform cell identification across multiple sessions. By comparing SCOUT with current popular cell tracking algorithms on simulated, 1-photon and 2-photon recordings, we empirically show this approach improves cell tracking quality, particularly when recordings exhibit spatial footprint movement between sessions or poor neural extraction quality.

Application of bioluminescence resonance energy transfer-based cell tracking approach in bone tissue engineering

Bioluminescent imaging (BLI) has emerged as a popular in vivo tracking modality in bone regeneration studies stemming from its clear advantages: non-invasive, real-time, and inexpensive. We recently adopted bioluminescence resonance energy transfer (BRET) principle to improve BLI cell tracking and generated the brightest bioluminescent signal known to date, which thus enables more sensitive real-time cell tracking at deep tissue level. In the present study, we brought BRET-based cell tracking strategy into the field of bone tissue engineering for the first time. We labeled rat mesenchymal stem cells (rMSCs) with our in-house BRET-based GpNLuc reporter and evaluated the cell tracking efficacy both in vitro and in vivo. In scaffold-free spheroid 3D culture system, using BRET-based GpNLuc labeling resulted in significantly better correlation to cell numbers than a fluorescence based approach. In scaffold-based 3D culture system, GpNLuc-rMSCs displayed robust bioluminescence signals with minimal background noise. Furthermore, a tight correlation between BLI signal and cell number highlighted the robust reliability of using BRET-based BLI. In calvarial critical sized defect model, robust signal and the consistency in cell survival evaluation collectively supported BRET-based GpNLuc labeling as a reliable approach for non-invasively tracking MSC. In summary, BRET-based GpNLuc labeling is a robust, reliable, and inexpensive real-time cell tracking method, which offers a promising direction for the technological innovation of BLI and even non-invasive tracking systems, in the field of bone tissue engineering.

Mesoscale Convective System Precipitation Characteristics over East Asia. Part I: Regional Differences and Seasonal Variations

AbstractMesoscale convective systems (MCSs) play an important role in modulating the global water cycle and energy balance and frequently generate high-impact weather events. The majority of existing literature studying MCS activity over East Asia is based on specific case studies and more climatological investigations revealing the precipitation characteristics of MCSs over eastern China are keenly needed. In this study, we use an iterative rain cell tracking method to identify and track MCS precipitation during 2008–16 to investigate regional differences and seasonal variations of MCS precipitation characteristics. Our results show that the middle-to-lower reaches of the Yangtze River basin (YRB-ML) receive the largest amount and exhibit the most pronounced seasonal cycle of MCS precipitation in eastern China. MCS precipitation over YRB-ML can exceed 2.6 mm day−1 in June, contributing over 30.0% of April–July total rainfall. Particularly long-lived MCSs occur over the eastern periphery of the Tibetan Plateau (ETP), with 25% of MCSs over the ETP persisting for more than 18 h in spring. In addition, spring MCSs feature larger rainfall areas, longer durations, and faster propagation speeds. Summer MCSs have a higher precipitation intensity and a more pronounced diurnal cycle except for southeastern China, where MCSs have similar precipitation intensity in spring and summer. There is less MCS precipitation in autumn, but an MCS precipitation center over the ETP still persists. MCSs reach peak hourly rainfall intensities during the time of maximum growth (a few hours after genesis), reach their maximum size around 5 h after genesis, and start decaying thereafter.

In vivo stem cell tracking using scintigraphy in a canine model of DMD

One of the main challenges in cell therapy for muscle diseases is to efficiently target the muscle. To address this issue and achieve better understanding of in vivo cell fate, we evaluated the relevance of a non-invasive cell tracking method in the Golden Retriever Muscular Dystrophy (GRMD) model, a well-recognised model of Duchenne Muscular Dystrophy (DMD). Mesoangioblasts were directly labelled with 111In-oxine, and injected through one of the femoral arteries. The scintigraphy images obtained provided the first quantitative mapping of the immediate biodistribution of mesoangioblasts in a large animal model of DMD. The results revealed that cells were trapped by the first capillary filters: the injected limb and the lung. During the days following injection, radioactivity was redistributed to the liver. In vitro studies, performed with the same cells prepared for injecting the animal, revealed prominent cell death and 111In release. In vivo, cell death resulted in 111In release into the vasculature that was taken up by the liver, resulting in a non-specific and non-cell-bound radioactive signal. Indirect labelling methods would be an attractive alternative to track cells on the mid- and long-term.

Effects of Freezing on Mesenchymal Stem Cells Labeled with Gold Nanoparticles.

Stem cell therapies are a promising treatment for many patients suffering from diseases with poor prognosis. However, clinical translation is inhibited by a lack of in vivo monitoring techniques to track stem cells throughout the course of treatment. Ultrasound-guided photoacoustic (PA) imaging of nanoparticle-labeled stem cells may be a solution. To allow PA tracking, stem cells must be labeled with an optically absorbing contrast agent. Gold nanoparticles are one option due to their cytocompatibility and strong optical absorption in the near-infrared region. However, stem cell labeling can require up to 24-h incubation with nanoparticles in culture before use. Although stem cell monitoring is critically needed, the additional preparation time may not be feasible-it is cost prohibitive and stem cell treatments should be readily available in emergency situations as well as scheduled procedures. To remedy this, stem cells can be labeled before freezing and long-term storage. While it is well known that stem cells retain their cellular function after freezing, storage, and thawing, the impact of gold nanoparticles on this process has yet to be investigated. Therefore, we assessed the viability, multipotency, and PA activity of gold nanosphere-labeled mesenchymal stem cells (MSCs) after freezing, storing, and thawing for 1 week, 1 month, or 2 months and compared to unlabeled, naive MSCs which were frozen, stored, and thawed at the same time points. Results indicated no substantial change in viability as assessed by the MTT assay. Differentiation, observed through adipogenesis and osteogenesis, was also comparable to controls. Finally, strong PA signals and similar PA spectral signatures remained. Further studies involving more diverse stem cell types and nanoparticles are required, but our data suggest that function and imaging properties of nanoparticle-labeled stem cells are maintained after freezing and storage, which improve translation of stem cell monitoring techniques by simplifying integration with clinical protocols. Impact statement Although stem cell tracking techniques are critically needed, stem cells must be labeled with contrast agents in advance of procedures, which is not clinically feasible due to increased procedure time. As a solution, a stock of labeled stem cells could be frozen and stored, ready for immediate use. Results showed that gold nanosphere-labeled stem cells can be frozen and stored long-term without impacting cellular function or photoacoustic imaging contrast, supporting further investigation of other contrast agents and cell types. Creating a bank of nanoparticle-labeled stem cells advances translation and scalability of stem cell tracking methods by improving integration with clinical protocols.

Tracking Superparamagnetic Iron Oxide-labeled Mesenchymal Stem Cells using MRI after Intranasal Delivery in a Traumatic Brain Injury Murine Model.

Stem cell-based therapies for brain injuries, such as traumatic brain injury (TBI), are a promising approach for clinical trials. However, technical hurdles such as invasive cell delivery and tracking with low transplantation efficiency remain challenges in translational stem-based therapy. This article describes an emerging technique for stem cell labeling and tracking based on the labeling of the mesenchymal stem cells (MSCs) with superparamagnetic iron oxide (SPIO)nanoparticles, as well as intranasal delivery of the labeled MSCs. These nanoparticles are fluorescein isothiocyanate (FITC)-embedded and safe to label the MSCs, which are subsequently delivered to the brains of TBI-induced mice by the intranasal route. They are then tracked non-invasively in vivo by real-time magnetic resonance imaging (MRI). Important advantages of this technique that combines SPIO for cell labeling and intranasal delivery include (1) non-invasive, in vivo MSC tracking after delivery for long tracking periods, (2) the possibility of multiple dosing regimens due to the non-invasive route of MSC delivery, and (3) possible applications to humans, owing to the safety of SPIO, non-invasive nature of the cell-tracking method by MRI, and route of administration.

Hyaluronan-Based Grafting Strategies for Liver Stem Cell Therapy and Tracking Methods.

Cell adhesion is essential for survival, it plays important roles in physiological cell functions, and it is an innovative target in regenerative medicine. Among the molecular interactions and the pathways triggered during cell adhesion, the binding of cluster of differentiation 44 (CD44), a cell-surface glycoprotein involved in cell-cell interactions, to hyaluronic acid (HA), a major component of the extracellular matrix, is a crucial step. Cell therapy has emerged as a promising treatment for advanced liver diseases; however, so far, it has led to low cell engraftment and limited cell repopulation of the target tissue. Currently, different strategies are under investigation to improve cell grafting in the liver, including the use of organic and inorganic biomatrices that mimic the microenvironment of the extracellular matrix. Hyaluronans, major components of stem cell niches, are attractive candidates for coating stem cells since they improve viability, proliferation, and engraftment in damaged livers. In this review, we will discuss the new strategies that have been adopted to improve cell grafting and track cells after transplantation.

A New Approach for Cell Detection and Tracking

In recent years, cell tracking methods by detection have become more and more popular because they outperformed cell tracking methods by contour evolution in most practical cell tracking applications. Yet, the most frequently used segmentation technique by cell detection methods is still threshold selection that is determined manually or by algorithms proposed in the 1970s. As a whole, these old threshold selection methods could not meet the accuracy requirement of cell detection adequately. In this paper, we propose a new approach of cell tracking by detection based on a multiple-threshold segmentation method that calculates multiple thresholds automatically and robustly. After cell detection, the proposed approach generates the timeline moving trajectory of a cell by connecting the cell positions along the time lapse image sequences based on morphological operations. We use four types of cells to verify the effectiveness of the proposed approach and the experimental results are favorable.

InVivo PET Tracking of 89Zr-Labeled Vγ9Vδ2T Cells to Mouse Xenograft Breast Tumors Activated with Liposomal Alendronate.

Gammadelta T (γδ-T) cells are strong candidates for adoptive immunotherapy in oncology due to their cytotoxicity, ease of expansion, and favorable safety profile. The development of γδ-T cell therapies would benefit from non-invasive cell-tracking methods and increased targeting to tumor sites. Here we report the use of [89Zr]Zr(oxinate)4 to track Vγ9Vδ2 T cells in vivo by positron emission tomography (PET). In vitro, we showed that 89Zr-labeled Vγ9Vδ2 T cells retained their viability, proliferative capacity, and anti-cancer cytotoxicity with minimal DNA damage for amounts of 89Zr ≤20 mBq/cell. Using a mouse xenograft model of human breast cancer, 89Zr-labeled γδ-T cells were tracked by PET imaging over 1 week. To increase tumor antigen expression, the mice were pre-treated with PEGylated liposomal alendronate. Liposomal alendronate, but not placebo liposomes or non-liposomal alendronate, significantly increased the 89Zr signal in the tumors, suggesting increased homing of γδ-T cells to the tumors. γδ-T cell trafficking to tumors occurred within 48 hr of administration. The presence of γδ-T cells in tumors, liver, and spleen was confirmed by histology. Our results demonstrate the suitability of [89Zr]Zr(oxinate)4 as a cell-labeling agent for therapeutic T cells and the potential benefits of liposomal bisphosphonate treatment before γδ-T cell administration.

Chronic TNFα-driven injury delays cell migration to villi in the intestinal epithelium.

The intestinal epithelium is a single layer of cells which provides the first line of defence of the intestinal mucosa to bacterial infection. Cohesion of this physical barrier is supported by renewal of epithelial stem cells, residing in invaginations called crypts, and by crypt cell migration onto protrusions called villi; dysregulation of such mechanisms may render the gut susceptible to chronic inflammation. The impact that excessive or misplaced epithelial cell death may have on villus cell migration is currently unknown. We integrated cell-tracking methods with computational models to determine how epithelial homeostasis is affected by acute and chronic TNFα-driven epithelial cell death. Parameter inference reveals that acute inflammatory cell death has a transient effect on epithelial cell dynamics, whereas cell death caused by chronic elevated TNFα causes a delay in the accumulation of labelled cells onto the villus compared to the control. Such a delay may be reproduced by using a cell-based model to simulate the dynamics of each cell in a crypt-villus geometry, showing that a prolonged increase in cell death slows the migration of cells from the crypt to the villus. This investigation highlights which injuries (acute or chronic) may be regenerated and which cause disruption of healthy epithelial homeostasis.

T-cell tracking using Cerenkov and radioluminescence imaging.

Cancer immunotherapy is a promising strategy based on the ability of the immune system to kill selected cells. In the development of an effective T-cell therapy, the noninvasive cell tracking methods play a crucial role. Here, we investigate the potentialities of T-cell marked with radionuclides in order to detect their localization with imaging techniques in small animal rodents. A protocol to label T-cells with 32 P-ATP was tested and evaluated. The homing of 32 P-ATP labeled T lymphocytes was investigated by Cerenkov luminescence imaging (CLI) and radioluminescence imaging (RLI). The first approach relies on the acquisition of Cerenkov photons produced by the beta particles emitted by the 32 P internalized by lymphocytes; the second one on the detection of photons coming from the conversion of radioactive energy in light done by scintillator crystals layered on the animals. The results show that T-cell biodistribution can be optically observed by both CLI and RLI in small animal rodents in in vivo and ex vivo acquisitions. T-cell localization in the tumor mass was definitively confirmed by flow cytometry.

PO-099 Targeting the mitogen activated protein kinase ERK5 in human melanoma

IntroductionPim kinases are constitutively active serine/threonine kinases that are upregulated by the JAK/STAT pathway and are often overexpressed in advanced human haematological and solid cancers. There they have been observed to enhance cancer cell growth, survival and energy metabolism. Using both cell-based and animal models, we have shown that PIM kinases also efficiently promote cancer cell migration, invasion and metastasis formation. In addition, we have identified several substrates to mediate the PIM-dependent effects.Material and methodsMotility of PC-3 prostate cancer cells was analysed by cell-based assays, such as wound healing assays with single cell tracking method, Boyden chamber assays and adhesion assays. The effects of PIM kinases on tumour growth were studied using subcutaneous and orthotopic mouse models for human prostate cancer. The interactions of PIM kinases and their substrates were investigated by in vitro kinase assays as well as microscopic techniques such as proximity ligation assay and fluorescence lifetime imaging. To reveal PIM-dependent functions, we used PIM-selective chemical inhibitors, transient or stable PIM upregulation as well as PIM silencing by RNA interference or CRISPR/Cas9-based gene editing.Results and discussionsPIM upregulation was shown to promote prostate cancer cell motility both in cell-based wound healing assays and in the orthotopically inoculated xenografts. By contrast, inhibition of PIM kinase activity by the pyrrolocarbazole compound DHPCC-9 decreased cell migration, invasion and adhesion as well as tumour growth, angiogenesis, lymphangiogenesis and formation of metastases. Furthermore, the PIM-dependent effects were shown to be mediated by the ability of PIM kinases to phosphorylate several key substrates, such as CXCR4, FOXP3, GSK3B, NFATC1 and NOTCH1. More recently, we have observed PIM kinases to influence also the actin cytoskeleton and have identified actin-regulating proteins as novel PIM substrates.ConclusionPIM kinases are promising targets for anti-cancer drug development, not only for their roles as pro-survival factors but also as pro-migratory factors. By inhibiting PIM kinases, it may be possible to simultaneously regulate many different PIM substrates that are essential for metastatic tumour growth.

PO-095 PIM kinases in the regulation of prostate cancer cell motility

Introduction Pim kinases are constitutively active serine/threonine kinases that are upregulated by the JAK/STAT pathway and are often overexpressed in advanced human haematological and solid cancers. There they have been observed to enhance cancer cell growth, survival and energy metabolism. Using both cell-based and animal models, we have shown that PIM kinases also efficiently promote cancer cell migration, invasion and metastasis formation. In addition, we have identified several substrates to mediate the PIM-dependent effects. Material and methods Motility of PC-3 prostate cancer cells was analysed by cell-based assays, such as wound healing assays with single cell tracking method, Boyden chamber assays and adhesion assays. The effects of PIM kinases on tumour growth were studied using subcutaneous and orthotopic mouse models for human prostate cancer. The interactions of PIM kinases and their substrates were investigated by in vitro kinase assays as well as microscopic techniques such as proximity ligation assay and fluorescence lifetime imaging. To reveal PIM-dependent functions, we used PIM-selective chemical inhibitors, transient or stable PIM upregulation as well as PIM silencing by RNA interference or CRISPR/Cas9-based gene editing. Results and discussions PIM upregulation was shown to promote prostate cancer cell motility both in cell-based wound healing assays and in the orthotopically inoculated xenografts. By contrast, inhibition of PIM kinase activity by the pyrrolocarbazole compound DHPCC-9 decreased cell migration, invasion and adhesion as well as tumour growth, angiogenesis, lymphangiogenesis and formation of metastases. Furthermore, the PIM-dependent effects were shown to be mediated by the ability of PIM kinases to phosphorylate several key substrates, such as CXCR4, FOXP3, GSK3B, NFATC1 and NOTCH1. More recently, we have observed PIM kinases to influence also the actin cytoskeleton and have identified actin-regulating proteins as novel PIM substrates. Conclusion PIM kinases are promising targets for anti-cancer drug development, not only for their roles as pro-survival factors but also as pro-migratory factors. By inhibiting PIM kinases, it may be possible to simultaneously regulate many different PIM substrates that are essential for metastatic tumour growth.

CGMP Signaling and Vascular Smooth Muscle Cell Plasticity.

Cyclic GMP regulates multiple cell types and functions of the cardiovascular system. This review summarizes the effects of cGMP on the growth and survival of vascular smooth muscle cells (VSMCs), which display remarkable phenotypic plasticity during the development of vascular diseases, such as atherosclerosis. Recent studies have shown that VSMCs contribute to the development of atherosclerotic plaques by clonal expansion and transdifferentiation to macrophage-like cells. VSMCs express a variety of cGMP generators and effectors, including NO-sensitive guanylyl cyclase (NO-GC) and cGMP-dependent protein kinase type I (cGKI), respectively. According to the traditional view, cGMP inhibits VSMC proliferation, but this concept has been challenged by recent findings supporting a stimulatory effect of the NO-cGMP-cGKI axis on VSMC growth. Here, we summarize the relevant studies with a focus on VSMC growth regulation by the NO-cGMP-cGKI pathway in cultured VSMCs and mouse models of atherosclerosis, restenosis, and angiogenesis. We discuss potential reasons for inconsistent results, such as the use of genetic versus pharmacological approaches and primary versus subcultured cells. We also explore how modern methods for cGMP imaging and cell tracking could help to improve our understanding of cGMP’s role in vascular plasticity. We present a revised model proposing that cGMP promotes phenotypic switching of contractile VSMCs to VSMC-derived plaque cells in atherosclerotic lesions. Regulation of vascular remodeling by cGMP is not only an interesting new therapeutic strategy, but could also result in side effects of clinically used cGMP-elevating drugs.

DETECTION AND TRACKING OF MIGRATING OLIGODENDROCYTE PROGENITOR CELLS FROM IN VIVO FLUORESCENCE TIME-LAPSE IMAGING DATA.

In this work, we develop a fully automatic algorithm named "MCDT" (Migrating Cell Detector and Tracker) for the integrated task of migrating cell detection, segmentation and tracking from in vivo fluorescence time-lapse microscopy imaging data. The interest of detecting and tracking migrating cells arouses from the scientific question in understanding the impact of oligodendrocyte progenitor cells (OPCs) migration in vivo, using advanced microscopy imaging techniques. Current practice of OPC mobility analysis relies on manual labeling, suffering from massive human labor, subjective biases, and weak reproducibility. Existing cell tracking methods have difficulties in analyzing such challenging data due to the extra complexity of in vivo data. Designed for in vivo data, MCDT circumvents the common strong assumption of separable feature distributions between foreground and background. Besides, by focusing on migrating cells (OPCs) only, MCDT relieves the burden of tracking all irrelevant cells correctly, not only accelerating the analysis but also achieving better accuracy in OPCs. Seed based segmentation and tracking by topology-preserved motion estimation endows MCDT with robustness to complex surroundings of the cell under tracking and to occasional inaccurate segmentation in some frames. We tested MCDT on imaging data of transgenic zebrafish larval spinal cord and MCDT showed very promising performance.