The ability to label and trace individual cells is a powerful experimental tool in many research areas including cell development, tumor evolution, stem cell differentiation, or carcinogenesis. It requires the ability to determine the location, distribution, and long-term viability of the cell populations as well as their biological fate with respect to cell activation and differentiation.
Molecular imaging techniques are avaible for the non-invasive monitoring of cellular dynamics in vivo.
NGS barcode labeling enables tracking of large numbers of clonal populations derived from individual founder cells.
Over the past decade, significant advances have been made in molecular imaging technology including bioluminescence imaging, fluorescence imaging, and enzyme-based positron emission tomography. Molecular imaging requires the selection of a molecular probe and imaging system. Protein probes are preferred, because proteins are more abundant within a cell.
BLI uses light generated from a luciferase enzyme-substrate and an ultrasensitive cooled charge-coupled camera for signal detection. BLI may have the highest imaging sensitivity (even down to the one cell level), is high-throughput, easy-to-use system, and low cost.
FLI uses red/green fluorescent protein (RFP or GFP) as a probe. Signal generation is achieved by exciting the fluorescent proteins at a given light wavelength and detection light emission at another wavelength with a charge-coupled camera. Compared to BLI, FLI is less sensitive with higher background. In most cases, FLI is used for live imaging of shallow tissues. RFP and GFP give more histological information and can be readily used for cell sorting. Now, FLI is normally coupled with BLI to provide an additional
means for cell selection, sorting, thus gaining more histological information.
Enzyme-based positron emission tomography uses HSV1-tk (Thymidine Kinase, TK) as a probe. Signal generation is achieved by the retention of radioisotope labeled chemicals for SPECT (single photon emission computed tomography) or PET imaging. This modality can be used for large animals or humans with detailed 3-dimensional capability but, it is more expensive to use this system.
Using CloneTracker NGS Barcode Libraries, it is possible to genomically label each cell with a uniquely identifiable short nucleotide sequence (i.e., a barcode). Since the barcodes genomically integrate, they are heritable so all progeny from each cell harbours the same unique sequence and clonal expansion for each founder cell can be monitored.
CloneTracker Expressed NGS Barcode Libraries enable tracking of clonal variations in large cell populations. When used in combination with single-cell RNA sequencing, they allow researchers to identify which genes are actually activated or shut down in different groups of cells so that, depending on the experiment, they can identify which genes are important for drug resistance, metastasis, cell differentiation, or other processes.