Difference between revisions of "Fluorescence Lifetime Imaging Microscopy Quantitative Measurements"
| Line 8: | Line 8: | ||
[[Fluorescence]] describes the emission of light by an atom or molecule that follows the absorption of electromagnetic energy [1]. When a fluorescent molecule absorbs energy, it is driven into an excited state that persists for a brief time. The molecule then transitions back to the lower energy ground state by one of several possible pathways. Some of the pathways for de-excitation to the ground state are illustrated in the simplified Perrin-Jabłoński plot in Figure 1. The pathways include internal conversion (<math>ic</math>), decay by fluorescence (<math>kf</math>), quenching (loss of excitation energy without the emission of light, <math>knf</math>), or intersystem crossing (<math>isc</math>) to the triplet state followed by decay by phosphorescence (<math>kpf</math>).The average time required for a population of fluorophores in the excited state to decay to the ground state is called the fluorescence lifetime, which is described by an exponential function (Figure 1): | [[Fluorescence]] describes the emission of light by an atom or molecule that follows the absorption of electromagnetic energy [1]. When a fluorescent molecule absorbs energy, it is driven into an excited state that persists for a brief time. The molecule then transitions back to the lower energy ground state by one of several possible pathways. Some of the pathways for de-excitation to the ground state are illustrated in the simplified Perrin-Jabłoński plot in Figure 1. The pathways include internal conversion (<math>ic</math>), decay by fluorescence (<math>kf</math>), quenching (loss of excitation energy without the emission of light, <math>knf</math>), or intersystem crossing (<math>isc</math>) to the triplet state followed by decay by phosphorescence (<math>kpf</math>).The average time required for a population of fluorophores in the excited state to decay to the ground state is called the fluorescence lifetime, which is described by an exponential function (Figure 1): | ||
| − | <math> I(t)=I_{0}e^{-t/\tau}\tag 1 | + | <math> I(t)=I_{0}e^{-t/\tau}\tag 1</math> |
where I(t) is the fluorescence impulse response at time t, I0 is the initial intensity after the excitation pulse, and τ is the fluorescence lifetime. | where I(t) is the fluorescence impulse response at time t, I0 is the initial intensity after the excitation pulse, and τ is the fluorescence lifetime. | ||
Revision as of 06:41, 16 February 2017
Department of Cellular and Integrative Physiology, Indiana University School of Medicine, 635 Barnhill Dr., Indianapolis, IN 46202 USA.
1. A Brief History of the Measurement of Fluorescence Lifetimes
Fluorescence describes the emission of light by an atom or molecule that follows the absorption of electromagnetic energy [1]. When a fluorescent molecule absorbs energy, it is driven into an excited state that persists for a brief time. The molecule then transitions back to the lower energy ground state by one of several possible pathways. Some of the pathways for de-excitation to the ground state are illustrated in the simplified Perrin-Jabłoński plot in Figure 1. The pathways include internal conversion ([math]ic[/math]), decay by fluorescence ([math]kf[/math]), quenching (loss of excitation energy without the emission of light, [math]knf[/math]), or intersystem crossing ([math]isc[/math]) to the triplet state followed by decay by phosphorescence ([math]kpf[/math]).The average time required for a population of fluorophores in the excited state to decay to the ground state is called the fluorescence lifetime, which is described by an exponential function (Figure 1):
[math] I(t)=I_{0}e^{-t/\tau}\tag 1[/math]
where I(t) is the fluorescence impulse response at time t, I0 is the initial intensity after the excitation pulse, and τ is the fluorescence lifetime.
