CellLight® Fluorescent Protein Constructs for Subcellular Structures
 | CellLight® reagents are ready-to-use fluorescent protein constructs targeted to specific subcellular structures. These reagents provide a simple and effective method for introducing targeted intracellular labels within living cells. Simply add the reagent to your cells, incubate overnight, and you’re ready to image your cells.
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Advantages of Labeling with CellLight® Reagents
- No lipids, chemical stains, or other potentially harmful treatments are required.
- Ready-to-use—simply add, incubate, then image
- No need to optimize transduction conditions more than once per cell type.
- Multiplex with cellular stains including other CellLight™ reagents, antibodies or MitoTracker® and LysoTracker® dyes.
- Efficient transduction of mammalian cells including "difficult" cells such as primary and stem cells.
CellLight® reagents come in a variety of colors and targets (Table 1), including actin, endsomes, lysosomes and tubulin, for convenient multiplexing and co-localization studies.
Overview of BacMam Delivery Technology
 | Created as ready-to-use BacMam expression and delivery reagents, the efficient transduction, robust expression and retained cellular function of these fusion proteins gives highly specific labeling for tools. Easily study dynamic changes, colocalization and signaling events in living cells. All CellLight® are compatible with fixed cell staining and high content analysis. A convenient BacMam 2.0 GFP Transduction Control is also available to easily determine how CellLight™ technology will work in your experimental system. For enhancement of the fluorescent protein signal in fixed samples, use one of our anti-GFP Antibodies.
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Table 1: CellLight® Reagents
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| CellLight® Reagents | CFP | GFP | RFP |
(440/480) | (488/510) | (555/584) | |
| Actin | |||
| Cytoplasm | |||
| Endoplasmic Reticulum | |||
| Early Endosomes | |||
| Late Endosomes | |||
| Golgi | |||
| Histones (Histone 2B) | |||
| Lysosomes | |||
| MAP4 | |||
| Mitochondria | |||
| Nucleus | |||
| Peroxisomes | |||
| Plasma Membrane | |||
| Synaptophysin | |||
| Talin | |||
| Tubulin | |||
| BacMam 2.0 Transduction Control | |||
| Null virus (control) | |||
Different cell types will have different transfection efficiencies with baculovirus -based technologies. Certain cell types, such as macrophages and other hematopoiteic cells, cannot be transfected with baculovirus. | |||
CellLight® Images
| CellLight® Golgi-GFP and CellLight® Mitochondria-RFP. Cascade Biologics® HASMC (human aortic smooth muscle cells) co-transduced with CellLight® Golgi-GFP and CellLight® Mitochondria-RFP and stained with Hoechst 33342. Imaging was performed on live cells using a Delta–Vision® Core microscope and standard DAPI/FITC/TRITC filter sets. |
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| CellLight® Talin-RFP and CellLight® Mitochondria-GFP. HeLa cells were transduced with CellLight® Talin-RFP and CellLight® Mitochondria-GFP and Hoechst 33342 (1 ug/ml). Imaging was performed on live cells using a Delta–Vision® Core microscope and standard DAPI/FITC/TRITC filter sets. |
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Real-time visualization of neuronal growth cone dynamics using BacMam 2.0
 | Live-cell imaging of neuronal growth cone dynamics in cultured rat hippocampal neurons. Hippocampal tissue from rat postnatal day 3 animals was harvested and dissociated by gentle trituration in neural culture medium prior to plating onto glial feeder cultures, which were established a week prior on 35 mm glass-bottomed dishes (MatTek®, Ashland, MA). Undissociated hippocampal tissue was allowed to settle, and the supernatant containing neurons was transferred to a fresh vial. Cell density was determined with the Countess® Automated Cell Counter, and cells were resuspended at 50,000 cells/mL in complete neural culture medium plus mitotic inhibitors. The medium was removed from the glial feeder cultures and replaced with 2 mL of the neural cell suspension. 50 µL each of BacMam 2.0 GFP transduction control and CellLight® MAP4-RFP were added to the dishes, and cells were placed into the cell culture incubator. A complete medium change was performed the following day, and cultures were allowed to mature for 2 weeks before imaging on a DeltaVison® DV Core (Applied Precision, Issaquah, WA), with partial media changes twice a week. GFP fluorescence was observed in neurons after 48 hr in culture. Images in the time-lapse series were collected with standard FITC and TRITC filter sets at 10-second intervals with 50-millisecond exposures. Images are a deconvoluted set of 10 projected optical sections 0.2 µm in depth. |
Time-lapse movie of U-2 OS cells transduced with CellLight® Histone 2B-RFP & CellLight® MAP4-GFP
 | U-2 OS cells were transduced with CellLight® Histone 2B-RFP and CellLight® MAP4-GFP and imaged the next day. Cells were imaged in McCoys media. Cells were kept in a climate controlled chamber and imaged with a 40X objective, images were collected every 5 minutes for 7 hours and 35 minutes. Fluorescence was detected using a Delta–Vision® Core microscope and the appropriate filters (FITC/TRITC). |
Time-lapse movie of HeLa cells transduced with CellLight® Mitochondria-GFP
 | HeLa cells were transduced with CellLight® Mitochondria-GFP and imaged the following day in MEM media with a 40X objective. Images were acquired at 5 second intervals over a period of 2 minutes and detected using the appropriate filters (FITC) on a Delta–Vision® Core microscope. |
