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Fluorescent proteins have revolutionized molecular and cellular biology, providing powerful tools for visualizing and understanding biological processes. These are generally naturally occurring proteins found in marine organisms, these fluorescent markers allow scientists to tag and track proteins, cells, and tissues with remarkable precision. Among the vast array of fluorescent proteins, several have become indispensable in research labs worldwide. This article sheds a spotlight on tdTomato and its peers.
The Foundation: Fluorescent Proteins
Fluorescent proteins emit light upon excitation by specific wavelengths, allowing visualization under fluorescence microscopy. The discovery of the green fluorescent protein (GFP) from the jellyfish Aequorea victoria paved the way for the development of various coloured fluorescent proteins through genetic engineering. These proteins have diverse applications, including studying gene expression, protein localization, and cell tracking.
Antibodies against these proteins enable precise detection, quantification, and analysis of these fluorescent proteins in various experimental contexts. Here is a detailed list of how antibodies specific to these fluorescent proteins are useful:
Name | Excitation (nm) | Emission (nm) | Color |
|---|---|---|---|
380 | 440 | ||
433 | 475 | ||
488 | 509 | ||
540 | 553 | ||
548 | 562 | ||
553 | 581 | ||
568 | 585 | ||
574 | 586 | ||
587 | 610 | ||
595 | 620 | ||
596 | 625 | ||
590 | 648 |
The Foundational Fluorescent Proteins
GFP (Green Fluorescent Protein), YFP (Yellow Fluorescent Protein), and RFP (Red Fluorescent Protein) are cornerstone tools in fluorescent imaging. These proteins are fundamental for visualizing and studying cellular and molecular processes in real time.
GFP (Green Fluorescent Protein)
GFP fluoresces green (excitation at 395/475 nm and emission at 509 nm) and has been extensively modified to improve its brightness and stability like eGFP (excitation at 484 nm and emission at 507nm).
YFP (Yellow Fluorescent Protein)
YFP a derivative of GFP, emits yellow fluorescence (excitation at 514 nm and emission at 527 nm) and is particularly useful in multicolour labelling experiments.
RFP (Red Fluorescent Protein)
RFP, with its far-red emission, is ideal for deep-tissue imaging and minimizing phototoxicity. These proteins are fundamental for visualizing and studying cellular and molecular processes in real time.
GFP, YFP, and RFP Antibody Applications
- Western Blotting: Detects GFP, YFP, and RFP-tagged proteins, validating their expression in various samples.
- Immunofluorescence: Localizes these tagged proteins within cells, aiding in the study of cellular dynamics and providing detailed imaging data.
- Flow Cytometry: Measures their expression in cell populations, facilitating cell sorting and functional analysis.
Compared to the hundreds of cyan, green, and yellow fluorescent proteins, only a few probes have been developed in the orange and red wavelengths (approximately 560 nm to 650 nm) of the spectrum. Nevertheless, the few that exist have been isolated from corals and show potential for use in a variety of imaging scenarios.
Probes commonly named red fluorescent protein RFP, such as DsRed, TagRFP, and tdTomato, actually have emission spectra that are significantly more orange than red. Regardless of the colour designation, orange spectral proteins are easier to image in multiple colours, such as cyan, green, and red, using standard tetramethylrhodamine isothiocyanate (TRITC) filter equipment.
tdTomato: The Bright Red Beacon
tdTomato is a standout red fluorescent protein known for its high brightness and photostability. Derived from a mutant of the Discosoma sp. red fluorescent protein (DsRed), tdTomato matures quickly, making it ideal for live-cell imaging and long-term studies. It emits a vivid red fluorescence (excitation at 554 nm and emission at 581 nm), which is easily distinguishable from other fluorescent proteins, facilitating multicolour imaging experiments.
The tandem dimer contains two copies of dTomato, linked by a 12-amino acid linker. Due to a pair of chromophores, tdTomato is extremely bright and has unique light stability. Reporter gene mice have been published, and it has no obvious toxicity to cells/mice, making them very suitable for live animal research. The biggest disadvantage of tdTomato is that it is a large molecule and may interfere with the folding of fusion proteins in some cases.
Researchers frequently use Biorbyt’s tdTomato antibody in live-cell imaging, multicolour labelling, transgenic animal models, gene expression studies, cell line development, and antibody-based detection techniques which are invaluable tools in the realm of life sciences.
tdTomato Antibody Applications
- Western Blotting: Detects tdTomato-tagged proteins, verifying their expression in cell or tissue lysates.
- Immunohistochemistry (IHC): Visualizes tdTomato expression in fixed tissues, providing spatial and morphological context.
- Immunoprecipitation (IP): Enriches tdTomato-tagged proteins from complex mixtures for further analysis.
mCherry: The Versatile Red Marker
mCherry is another popular red fluorescent protein derived from DsRed. It exhibits excitation at 587 nm and emission at 610 nm, providing bright and stable red fluorescence. mCherry is monomeric, making it suitable for fusion with other proteins without disrupting their function. Its robustness and versatility have made it a go-to marker for various applications, including studying protein-protein interactions and monitoring cellular events.
mCherry Antibody Applications
- Western Blotting: Detects mCherry-tagged proteins, confirming their expression levels.
- Immunofluorescence (IF): Localizes mCherry-tagged proteins within cells, revealing subcellular structures and dynamics.
- Flow Cytometry: Quantifies mCherry expression in cell populations, useful for sorting and phenotyping.
mOrange: The Bright Orange Option
mOrange is derived from DsRed and offers bright orange fluorescence (excitation at 548 nm and emission at 562 nm). Its distinct colour makes it valuable for multicolour labelling experiments, allowing researchers to visualize multiple cellular components simultaneously. mOrange's brightness and stability have made it a favourite in studies requiring precise and clear imaging.
mOrange Antibody Applications
- Western Blotting: Detects mOrange-tagged proteins, confirming their expression profiles.
- Immunofluorescence (IF):Provides visualization of mOrange-tagged proteins in various cell types and tissues.
- Immunoprecipitation: Isolates mOrange-tagged proteins from complex biological samples.
mScarlet: The Bright Red Innovator
mScarlet is a newer addition to the family of red fluorescent proteins, engineered for exceptional brightness and maturation speed. With excitation at 569 nm and emission at 594 nm, mScarlet outshines many other red fluorescent proteins. Its high quantum yield and photostability make it an excellent choice for super-resolution microscopy and other advanced imaging techniques.
mScarlet Antibody Applications
- Western Blotting: Detects mScarlet-tagged proteins, validating their presence and expression.
- Immunofluorescence (IF): Visualizes mScarlet-tagged proteins within cellular contexts, providing insights into cellular architecture
- Co-immunoprecipitation (Co-IP): Studies interactions between mScarlet-tagged proteins and their partners.
Biorbyt offers a wide range of high-quality antibodies for popular fluorescent proteins. Our antibodies are designed for optimal specificity and sensitivity, suitable for applications like Western blotting, immunofluorescence, flow cytometry, immunohistochemistry, and immunoprecipitation. Enhance your research with our reliable antibodies, tailored to deliver precise and reproducible results.