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The only way to prove that a system is as good as you say is to show the actual data
When we developed the EasyLife™ TCSPC, we knew that some people would be skeptical that an instrument that is so inexpensive, and so small, and so easy to operate, could possibly provide research quality results. That is why are compiling a growing collection of data acquired on the EasyLife™ TCSPC. Please review the links below for to see for yourself just how good the system can work for you.
Our experience in lifetime instrumentation development goes back almost 40 years to the introduction of the first commercial TCSPC instrument. We have been well aware of the many potential uses of this technique. In the past, the barrier for the acceptance of fluorescence lifetimes has been the high price and the complexity of the equipment. EasyLife™ TCSPC is a major breakthrough that has changed all that. Now the EasyLife™ TCSPC can truly be used in research labs that previously could not afford the technology, or did not have the experience to use the technique.
Applications for the EasyLife™
Click on the application for more information.
Protein Folding/Unfolding
Most proteins fluoresce due to the presence of any or all three fluorescent amino acids: tryptophan, tyrosine, and phenylalanine. Intrinsic time-resolved fluorescence of tryptophan is commonly used to study the structure and dynamics of proteins. These experiments require pulsed light sources emitting in the UV, between 270 and 295 nm. The EasyLifeT TCSPC, equipped with the 280 or 295 nm pulsed LED source, is a very robust yet fast instrument perfectly suited for use with tryptophan and tyrosine fluorophores.
If you happen to use external fluorophores, there is a large selection of pulsed LEDs available for any wavelength in the UV-VIS range. A polarity sensitive, hydrophobic probe such as ANS is a good illustration of binding of an extrinsic probe to a protein. ANS binding to bovine serum albumin BSA is easily monitored with the EasyLifeT TCSPC equipped with the 370 nm LED. The lifetime of ANS in the buffer is very short and it increases upon binding to BSA. The ratio of free ANS to BSA bound ANS can be easily determined from the double exponential fit to the fluorescence decay.
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Fluorescence decays of bovine serum albumin (BSA) in PBS buffer were measured with the EasyLifeT TCSPC (295nm excitation LED). The native protein shows a complex triple-exponential decay with an increasing prevalence of the longer lived components. After being treated with SDS detergent, BSA undergoes a structural transition and its fluorescence decay shifts to shorter lifetimes showing again a triple-exponential decay with an increased prevalence of the shorter components. |
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DNA (Nucleic Acid)
If you study conformational features or hybridization of DNA, the EasyLifeT TCSPC is the right system for you. A probe molecule in a buffer will show very little or no anisotropy. Attach it to a protein, DNA, or membrane, however, and the anisotropy is increased. This is all that the steady state experiment can tell you: the probe is attached to a much bigger entity. However, if you measure the lifetime of the probe, you can estimate the rate of rotational diffusion in addition to the size of the macromolecule that is attached to the probe.
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Ethidium Bromide (EB) is a commonly used DNA probe, which readily intercalates between the DNA bases. EB (excitation 460nm, emission 580nm) is weakly fluorescent in aqueous media, but becomes strongly fluorescent after intercalation into DNA. The lifetime of EB in buffer is 1.71 ns and increases dramatically to 22.7 ns after binding to calf thymus DNA. |
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POPOP
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BSA Protein Rotational Correlation Times
Time resolved fluorescence anisotropy (polarization) is a powerful experimental tool. A probe molecule in a buffer will show no, or very little, anisotropy. Attach it to a protein, DNA, membrane etc. and the anisotropy is increased. This is all that the steady state experiment can tell you: the probe is attached to a much bigger entity. However, if you measure the lifetime of the probe, you can estimate the rate of rotational diffusion and the size of the macromolecule your probe is attached to.
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Short Lifetimes with ps Laser Diodes
One inherent benefit of the TCSPC detection technique is the ability to measure shorter lifetimes than is possible with the EasyLife™ V using the stroboscopic detection technique. With the EasyLife™ TCSPC you can take advantage of the newest picosecond laser diodes and detect shorter fluorescence lifetimes. The combination of the two can make for a very powerful system. Contact OBB for a list of available picosecond laser diodes.
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For more information on the EasyLife™ TCSPC send us a quote request or contact OBB!
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