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Fluorescent Sensor
Overview of Fluorescent Sensor
Biological sensors are fundamental to current research, from quantifying metabolites under different perturbations to identifying cell states. Many types of sensors exist with a variety of readouts, with one of the most widely used platforms being fluorescent sensors. Fluorescent sensor molecules for imaging cellular molecules have become a hot topic in chemical biology research during the last two decades. A fluorescent sensor is advantageous due to its high sensitivity.
Fig.1 Metabolic targets of fluorescent biosensors in neurons.1
Services at Creative Biolabs
With professional expertise, advanced technology platforms, excellent specialists, and rich experience in neuroscience, Creative Biolabs has gained a good reputation in the industry. Years of practice devoted to neuroscience research make us experienced in fluorescent sensors' application in this field. With all these advantages, we are confident in providing customers high-quality relevantly custom productions. If you are interested in applying fluorescent sensors in neuroscience research or have any other inquiries on neuro-based custom productions, please feel free to contact us for more information.
Our services cater to researchers and scientists studying various aspects of the brain and nervous system.
- Our services include the design, development, and production of fluorescent sensors that can be used to monitor specific biomolecules or physiological parameters in real-time. These sensors are engineered to emit fluorescence signals when they bind to their target molecules, providing researchers with valuable insights into cellular processes and activities.
- Our team of experts works closely with clients to customize fluorescent sensors based on their specific research needs. We offer a wide range of sensor options, including calcium sensors, pH sensors, neurotransmitter sensors, and ion sensors, among others. These sensors can be delivered in various formats, such as genetically encoded sensors for use in cell culture and animal models, as well as small molecule probes for in vitro studies.
- Our services also include validation, optimization, and troubleshooting support to ensure the success of your experiments. We provide comprehensive training and guidance to help researchers integrate our sensors into their experimental workflows effectively.
Our services are designed to empower neuroscience researchers with advanced tools and technologies. We also offer other related services, including but not limited to:
| Services | Descriptions |
|---|---|
| Calcium Assay | We offer novel calcium assays to clients around the world, including customized simple and quantitative methods for measuring calcium in different biological fluids. |
| Optogenetic Indicators | We provide optogenetic tools, including a variety of optogenetic indicators that can be easily delivered to target neuronal populations, using a variety of genetic approaches to achieve cell type-specific manipulation. |
| STEMOD™ Advanced Drug Discovery | We develop integrated technology platforms to provide one-stop CNS drug discovery services, including studies on BBB transport and distribution in the brain. |
Published Data
The monoamine neuromodulator dopamine (DA) plays a crucial role in the brain, and the ability to directly measure dopaminergic activity is essential for understanding its physiological function. Therefore, Fangmiao Sun et al. developed the first DA sensor based on red fluorescent GPCR activation and an optimized version of green fluorescent DA sensor.
Both the red and green DA sensors have substantial fluorescence increases with subcellular resolution, subsecond kinetics and nanomolar to submicromolar affinity. The results indicate that these sensors can be used to measure DA release in mouse brain slices, as well as in the Drosophila olfactory system and in the NAc of freely moving mice during sexual behavior. Importantly, these sensors can report DA release induced by electrical stimulation, optogenetic activation, and a variety of physiologically relevant stimuli and behaviors.
Fig. 2 Development and characterization of two novel red fluorescent DA sensors and second-generation green fluorescent DA sensors.2
Application of Fluorescent Sensor in Neuroscience
In neuroscience, the most applied fluorescent sensors are genetically encoded calcium indicators (GECIs), genetically encoded voltage indicators (GEVIs), and pH sensors (pHluorins). Numerous other fluorescent indicators have been applied to study brain cell activity, including sensors for inorganic ions and molecules (chloride, zinc, hydrogen peroxide) and organic signaling molecules and metabolites (glutamate, acetylcholine, ATP, NADH, cyclic nucleotides, glucose, pyruvate, phospholipids). Several sensors that directly monitor enzyme activity have also been developed. These allow the activity of small GTPases, kinases (e.g., protein kinase A), and proteases to be detected. Moreover, GPCR activation can be followed by intra- or intermolecular FRET, either within the heterodimeric G protein or between the GPCR and a binding partner (G protein or arrestin). This approach has been applied to analyze the activation kinetics of metabotropic glutamate receptors, GABAB receptors, and M1 muscarinic acetylcholine receptors.
| Sensor | Scaffold | Fluorescent Protein(s) | Excitation | Emission | Sensor Design | Dynamic range: fold change or Δ lifetime | Affinity (Kd or KR) |
|---|---|---|---|---|---|---|---|
| ATP | |||||||
| ATeam1.03 | F0F1-ATP synthase, ε subunit (B. subtilis) | mseCFP/ mVenus | 435 nm (D) |
475 nm (D) 527 nm (A) |
FRET | 2.3-fold (37℃) | 3.3 mM |
| ATeam1.03YEMK | F0F1-ATP synthase, ε subunit (B. subtilis) | mseCFP/ mVenus | 435 nm (D) |
475 nm (D) 527 nm (A) |
FRET | n.r. |
1.2 mM (37℃) 2.6 mM (20-22℃) |
| QUEEN-2m | F0F1-ATP synthase, ε subunit (B. subtilis) | cp-EGFP | 400 nm / 494 nm | 513 nm | Ratiometric (excitation) | >3-fold (25℃) | 2.4 mM |
| QUEEN-7μ | F0F1-ATP synthase, ε subunit (B. PS3) | cp-EGFP | 400 nm / 494 nm | 513 nm | Ratiometric (excitation) |
~4.3-fold (37℃) ~5-fold (25℃) |
14 μM 7.2 μM |
| iATPSnFR1.0 | F0F1-ATP synthase, ε subunit (B. PS3) | cp-SFGFP | 488 nm | 515 nm | Intensity | 2-fold (RT) | 350 μM |
| iATPSnFR1.1 | F0F1-ATP synthase, ε subunit (B. PS3) | cp-SFGFP | 488 nm | 515 nm | Intensity | 1.88-fold (RT) | 138 μM |
| ATP:ADP | |||||||
| PercevalHR | GlnK, nucleotide binding protein (M. jannaschii) | cp-mVenus | 482 nm / 455 nm | 529 nm | Ratiometric (excitation) |
4.6-fold (37℃) ~4-fold (RT) |
ATP:ADP ≈ 6.1 ATP:ADP ≈ 3.5 |
| NADH | |||||||
| Frex | B-Rex, NADH binding protein (B. subtilis) | cpYFP | 488 nm / 405 nm | 525 nm | Ratiometric (excitation) | ~9.5-fold (RT) | 3.7 μM |
| NADH:NAD+ | |||||||
| SoNar | T-Rex, NADH binding protein (T. aquaticus) | cpYFP | 420 nm / 485 nm | 528 nm | Ratiometric (excitation) | ~15-fold (RT) | NADH:NAD+ ≈ 1/40 |
| Peredox | T-Rex, NADH binding protein (T. aquaticus) | cp-T- Sapphire | 400 nm | 510 nm | Intensityb | 2.5-fold (35℃) | NADH:NAD+ ≈ 1/90 |
| 800 nm (two-photon) | 525 nm | Lifetime |
0.9 ns (35℃) 0.8 ns (25℃) |
NADH:NAD+ ≈ 1/255 NADH:NAD+ ≈ 1/529 | |||
| Glucose | |||||||
| FLII12Pglu700μ∆6 | MglB, glucose/galactose binding protein (E. coli) | eCFP/ Citrine | 433 nm (D) |
485 nm (D) 528 nm (A) |
FRET | 1.5-fold (RT) | 660 μM |
| Green Glifon600 | MglB, glucose/galactose binding protein (E. coli) | Citrine | 480 nm | 530 nm | Intensity | ~5-fold (RT) | 590 μM |
| Green Glifon4000 | MglB, glucose/galactose binding protein (E. coli) | Citrine | 480 nm | 530 nm | Intensity | ~6-fold (RT) | 3.8 mM |
| iGlucoSnFR | GGBP, glucose/ galactose binding protein (T. thermophilus) | cpGFP | 485 nm | 515 nm | Intensity | 3.32-fold (RT) | 7.7 mM |
| iGlucoSnFR-TS | GGBP, glucose/ galactose binding protein (T. thermophilus) | cp-T- Sapphire | 790 nm (two-photon) | 525 nm | Lifetime |
0.34 ns (37℃) 0.38 ns (37℃) |
2.2 mM 1.8 mMc |
| Lactate | |||||||
| Laconic | LldR, lactate binding transcription regulator (E. coli) | mTFP/ Venus | 430 nm (D) |
480 nm (D) 535 nm (A) |
FRET | ~1.2-fold (25℃) |
Biphasic: K1 = 8 µM K2 = 830 µM |
| Pyruvate | |||||||
| Pyronic | PdhR, pyruvate dehydrogenase complex repressor (E. coli) | mTFP/ Venus | 430 nm (D) |
480 nm (D) 535 nm (A) |
FRET | ~1.24-fold (RT) | 107 μM |
Table1 Genetically encoded fluorescent biosensors used for the study of neuronal metabolism
FAQs
Q: Can you help with the optimization of fluorescent sensors for specific brain regions or cell types?
A: Yes, we provide comprehensive support in optimizing fluorescent sensors for specific brain regions or cell types. Our team of experts collaborates with you to select the most suitable sensors and optimize their expression and functionality. We can assist with targeted delivery systems, such as viral vectors, and fine-tune sensor parameters like sensitivity, dynamic range, and brightness to ensure reliable performance in your desired neural circuits.
Q: What kind of support do you offer for integrating your fluorescent sensors into my experimental setup?
A: We offer extensive support for integrating our fluorescent sensors into your experimental setup, from sensor selection to data analysis. Our team can assist with experimental design, including choosing the right sensors, imaging systems, and data acquisition methods. We also provide protocols and troubleshooting support to help you achieve consistent results. Additionally, we offer training and consultation services to ensure that your lab is fully equipped to handle sensor-based experiments.
Q: Are your fluorescent sensors compatible with different imaging platforms?
A: Our fluorescent sensors are compatible with a variety of imaging platforms, including widefield microscopy, confocal microscopy, and two-photon imaging. We also offer sensors optimized for advanced techniques like optogenetics and high-speed imaging. Whether you are working with in vitro cultures or performing in vivo imaging in animal models, our sensors can be adapted to suit your platform, ensuring high-resolution and high-sensitivity measurements of neural activity.
Q: How do you ensure the specificity and sensitivity of fluorescent sensors for different neurotransmitters?
A: Our fluorescent sensors are meticulously engineered for high specificity and sensitivity to different neurotransmitters. We achieve this through advanced protein engineering and molecular optimization techniques. For example, our neurotransmitter sensors are designed with binding domains that selectively interact with specific neurotransmitters while minimizing cross-reactivity. We also validate each sensor's performance across various conditions, ensuring that they deliver precise and consistent signals for accurate neurotransmitter detection in both in vitro and in vivo experiments.
Scientific Resources
References
- Koveal, Dorothy, Carlos Manlio Díaz-García, and Gary Yellen. "Fluorescent biosensors for neuronal metabolism and the challenges of quantitation." Current Opinion in Neurobiology 63 (2020): 111-121.
- Sun, Fangmiao, et al. "New and improved GRAB fluorescent sensors for monitoring dopaminergic activity in vivo." BioRxiv (2020): 2020-03.
For Research Use Only. Not For Clinical Use.