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The Development of Fluorescence Intensity Standards

View Article: PubMed Central - PubMed

ABSTRACT

The use of fluorescence as an analytical technique has been growing over the last 20 years. A major factor in inhibiting more rapid growth has been the inability to make comparable fluorescence intensity measurements across laboratories. NIST recognizes the need to develop and provide primary fluorescence intensity standard (FIS) reference materials to the scientific and technical communities involved in these assays. The critical component of the effort will be the cooperation between the Federal laboratories, the manufacturers, and the technical personnel who will use the fluorescence intensity standards. We realize that the development and use of FIS will have to overcome many difficulties. However, as we outline in this article, the development of FIS is feasible.

No MeSH data available.


A schematic of the geometry of illumination and detection. The shaded volume is the sensing volume and is the volume over which the integral in Eq. (2) is taken. The sensing volume is the overlap of the illumination volume bounded by I and I′ and the detection volume bounded by D, D′. The geometry and various filters and optical elements give the instrumental factors in the fluorescence intensity measurements as described in Eq. (3).
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f3-j62gai: A schematic of the geometry of illumination and detection. The shaded volume is the sensing volume and is the volume over which the integral in Eq. (2) is taken. The sensing volume is the overlap of the illumination volume bounded by I and I′ and the detection volume bounded by D, D′. The geometry and various filters and optical elements give the instrumental factors in the fluorescence intensity measurements as described in Eq. (3).

Mentions: Given a source of photons with known energies (spectral characteristics), the excitation optics will select a set of wavelengths that are incident on the sample. The excitation optics will illuminate a specific volume inside the sample (bounded by I and I′ in Fig. 3). The collection optics will collect light emitted from a volume element that we call the detection volume (bounded by D and D′ in Fig. 3). The total fluorescence intensity after the collection optics will depend on the overlap of the illumination and detection volumes and we call these overlapping volumes the sensing volume. Figure 3 is a schematic representation of the sensing volume. Only fluorophore molecules in the sensing volume can contribute to the fluorescence signal.


The Development of Fluorescence Intensity Standards
A schematic of the geometry of illumination and detection. The shaded volume is the sensing volume and is the volume over which the integral in Eq. (2) is taken. The sensing volume is the overlap of the illumination volume bounded by I and I′ and the detection volume bounded by D, D′. The geometry and various filters and optical elements give the instrumental factors in the fluorescence intensity measurements as described in Eq. (3).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4862808&req=5

f3-j62gai: A schematic of the geometry of illumination and detection. The shaded volume is the sensing volume and is the volume over which the integral in Eq. (2) is taken. The sensing volume is the overlap of the illumination volume bounded by I and I′ and the detection volume bounded by D, D′. The geometry and various filters and optical elements give the instrumental factors in the fluorescence intensity measurements as described in Eq. (3).
Mentions: Given a source of photons with known energies (spectral characteristics), the excitation optics will select a set of wavelengths that are incident on the sample. The excitation optics will illuminate a specific volume inside the sample (bounded by I and I′ in Fig. 3). The collection optics will collect light emitted from a volume element that we call the detection volume (bounded by D and D′ in Fig. 3). The total fluorescence intensity after the collection optics will depend on the overlap of the illumination and detection volumes and we call these overlapping volumes the sensing volume. Figure 3 is a schematic representation of the sensing volume. Only fluorophore molecules in the sensing volume can contribute to the fluorescence signal.

View Article: PubMed Central - PubMed

ABSTRACT

The use of fluorescence as an analytical technique has been growing over the last 20 years. A major factor in inhibiting more rapid growth has been the inability to make comparable fluorescence intensity measurements across laboratories. NIST recognizes the need to develop and provide primary fluorescence intensity standard (FIS) reference materials to the scientific and technical communities involved in these assays. The critical component of the effort will be the cooperation between the Federal laboratories, the manufacturers, and the technical personnel who will use the fluorescence intensity standards. We realize that the development and use of FIS will have to overcome many difficulties. However, as we outline in this article, the development of FIS is feasible.

No MeSH data available.