Edward A. Early

Quantitating Fluorescence Intensity From Fluorophore: The Definition of MESF Assignment

The quantitation of fluorescence radiance may at first suggest the need to obtain the number of fluorophore that are responsible for the measured fluorescence radiance. This goal is beset by many difficulties since the fluorescence radiance depends on three parameters 1) the probability of absorbing a photon (molar extinction), 2) the number of fluorophores, and 3) the probability of radiative decay of the excited state (quantum yield). If we use the same fluorophore in the reference solution and the analyte then, to a good approximation, the molar extinction drops out from the comparison of fluorescence radiance and we are left with the comparison of fluorescence yield which is defined as the product of fluorophore concentration and the molecular quantum yield. The equality of fluorescence yields from two solutions leads to the notion of equivalent number of fluorophores in the two solutions that is the basis for assignment of MESF (Molecules of Equivalent Soluble Fluorophore) values. We discuss how MESF values are assigned to labeled microbeads and by extension to labeled antibodies, and how these assignments can lead to the estimate of the number of bound antibodies in flow cytometer measurements.

Bidirectional reflectance round-robin in support of the Earth Observing System program

Abstract Laboratory measurements of the bidirectional reflectance distribution function (BRDF) of diffuse reflectors are required to support calibration in the Earth Observing System (EOS) program of the National Aeronautics and Space Administration. To assess the ability of the instrument calibration laboratories to perform accurate BRDF measurements, a round-robin with the National Institute of Standards and Technology (NIST) as the central laboratory was initiated by the EOS Project Science Office. The round-robin parameters include sample type, wavelength, and incident and viewing angles. The results show that the participating calibration laboratories are, with a few exceptions due to experimental techniques or sample properties, generally able to measure BRDF for the round-robin parameters to within 2% of the values measured by NIST.

The 1996 North American Interagency Intercomparison of Ultraviolet Monitoring Spectroradiometers

Concern over stratospheric ozone depletion has prompted several government agencies in North America to establish networks of spectroradiometers for monitoring solar ultraviolet irradiance at the surface of the Earth. To assess the ability of spectroradiometers to accurately measure solar ultraviolet irradiance, and to compare the results between instruments of different monitoring networks, the first North American Intercomparison of Ultraviolet Monitoring Spectroradiometers was held September 19–29, 1994 at Table Mountain outside Boulder, Colorado, USA. This Intercomparison was coordinated by the National Institute of Standards and Technology and the National Oceanic and Atmospheric Administration (NOAA). Participating agencies were the Environmental Protection Agency, National Science Foundation, Smithsonian Environmental Research Center, and Atmospheric Environment Service, Canada. Instruments were characterized for wavelength accuracy, bandwidth, stray-light rejection, and spectral irradiance responsivity, the latter with a NIST standard lamp calibrated to operate in the horizontal position. The spectral irradiance responsivity was determined once indoors and twice outdoors, and demonstrated that, while the responsivities changed upon moving the instruments, they were relatively stable when the instruments remained outdoors. Synchronized spectral scans of the solar irradiance were performed over several days. Using the spectral irradiance responsivities determined with the NIST standard lamp, and a simple convolution technique to account for the different bandwidths of the instruments, the measured solar irradiances agreed within 5 %.

Methodology for Deriving Clear-Sky Erythemal Calibration Factors for UV Broadband Radiometers of the U.S. Central UV Calibration Facility

Abstract In the United States, there are several federal agencies interested in the effects of UV radiation, which has resulted in the establishment of UV monitoring programs each with their own instrumentation and sites designed to address their specific needs. In 1993, participating agencies of the U.S. Global Change Research Program organized a UV Panel for coordinating the different agencies’ programs in order to ensure that UV data are intercalibrated, have common quality assurance and control procedures, and that the efforts among agencies are not duplicated. In order to achieve these goals, in 1994 the UV Panel recommended formation of the U.S. Central UV Calibration Facility (CUCF), which is operated by the Surface Radiation and Research Branch of the Air Resources Laboratory of National and Oceanic Atmospheric Administration. The CUCF is responsible for characterizing and calibrating UV measuring instruments from several U.S. federal agencies. Part of this effort is to calibrate UVB broadband rad...

The 1997 North American Interagency Intercomparison of Ultraviolet Spectroradiometers Including Narrowband Filter Radiometers

The fourth North American Intercomparison of Ultraviolet Monitoring Spectroradiometers was held September 15 to 25, 1997 at Table Mountain outside of Boulder, Colorado, USA. Concern over stratospheric ozone depletion has prompted several government agencies in North America to establish networks of spectroradiometers for monitoring solar ultraviolet irradiance at the surface of the Earth. The main purpose of the Intercomparison was to assess the ability of spectroradiometers to accurately measure solar ultraviolet irradiance, and to compare the results between instruments of different monitoring networks. This Intercomparison was coordinated by NIST and NOAA, and included participants from the ASRC, EPA, NIST, NSF, SERC, USDA, and YES. The UV measuring instruments included scanning spectroradiometers, spectrographs, narrow band multi-filter radiometers, and broadband radiometers. Instruments were characterized for wavelength accuracy, bandwidth, stray-light rejection, and spectral irradiance responsivity. The spectral irradiance responsivity was determined two to three times outdoors to assess temporal stability. Synchronized spectral scans of the solar irradiance were performed over several days. Using the spectral irradiance responsivities determined with the NIST traceable standard lamp, and a simple convolution technique with a Gaussian slit-scattering function to account for the different bandwidths of the instruments, the measured solar irradiance from the spectroradiometers excluding the filter radiometers at 16.5 h UTC had a relative standard deviation of ±4 % for wavelengths greater than 305 nm. The relative standard deviation for the solar irradiance at 16.5 h UTC including the filter radiometer was ±4 % for filter functions above 300 nm.

Intrinsic Wavelength Standard Absorption Bands in Holmium Oxide Solution for UV/visible Molecular Absorption Spectrophotometry

The transmittance minima of 18 absorption bands of a solution of 40 g/L holmium oxide in 10% (volume fraction) perchloric acid are certified as intrinsic traceable wavelength standards, by means of a multicenter measurement on material from a single source coupled with comparisons of a variety of preparations of the material evaluated on a single instrument. Fit-for-purpose artifact standards for the experimental calibration or validation of wavelength scales of chemical spectrophotometers can be carefully produced by end users themselves or by commercial standards producers. The intrinsic (data) standard confers traceability to the SI unit of length in place of costly transfer artifacts and repetitive calibration procedures. Certified values are provided for instrumental spectral bandwidths of 0.1–3.0 nm in 0.1 nm intervals, and information values are provided to a spectral bandwidth of 10 nm at wider intervals. Expanded uncertainties are typically less than ±0.1 nm for certified band positions.

Irradiance of Horizontal Quartz-Halogen Standard Lamps

Spectral irradiance calibrations often require that irradiance standard lamps be oriented differently than the normal calibration orientation used at the National Institute of Standards and Technology and at other standards laboratories. For example, in solar measurements the instruments are generally upward viewing, requiring horizontal working standards for minimization of irradiance calibration uncertainties. To develop a working standard for use in a solar ultraviolet intercomparison, NIST determined the irradiance of quartz-halogen lamps operating in the horizontal position, rather than in the customary vertical position. An experimental technique was developed which relied upon equivalent lamps with independent mounts for each orientation and a spectroradiometer with an integrating sphere whose entrance port could be rotated 90° to view either lamp position. The results presented here are limited to 1000 W quartz-halogen type lamps at ultraviolet wavelengths from 280 nm to 400 nm. Sources of uncertainty arose from the lamps, the spectroradiometer, and the lamp alignment, and increased the uncertainty in the irradiance of horizontal lamps by less than a factor of two from that of vertical NIST standard lamps. The irradiance of horizontal lamps was less than that of vertical lamps by approximately 6 % at long wavelengths (400 nm) to as much as 12 % at the shortest wavelengths (280 nm). Using the Wien radiation law, this corresponds to color temperature differences of 15.7 K and 21.3 K for lamps with clear and frosted envelopes, respectively.

ULTRAVIOLET SPECTRAL IRRADIANCE SCALE COMPARISON : 210 NM TO 300 NM

Comparison of the irradiances from a number of ultraviolet spectral irradiance standards, based on different physical principles, showed agreement to within their combined standard uncertainties as assigned to them by NIST. The wavelength region of the spectral irradiance comparison was from 210 nm to 300 nm. The spectral irradiance sources were: an electron storage ring, 1000 W quartz-halogen lamps, deuterium arc lamps, and a windowless argon miniarc.

Radiometric calibration of the Scripps Earth Polychromatic Imaging Camera

As part of the Triana mission, the Scripps Earth Polychromatic Imaging Camera (Scripps-EPIC) will view the full sunlit side of Earth from the Lagrange-1 point. The National Institute of Standards and Technology and the Scripps Institution of Oceanography, in collaboration with the contractor, Lockheed-Martin, planned the radiometric calibration of Scripps-EPIC. The measurements for this radiometric calibration were selected based upon the optical characteristics of Scripps-EPIC, the measurement equation relating signal to spectral radiance, and the available optical sources and calibrated radiometers. The guiding principle for the calibration was to perform separate, controlled measurements for each parameter in the measurement equation, namely dark signal, linearity, exposure time, and spectral radiance responsivity.

Standard Reference Material 2036 Near-Infrared Reflection Wavelength Standard

Standard Reference Material 2036 (SRM 2036) is a certified transfer standard intended for the verification and calibration of the wavelength/wavenumber scale of near-infrared (NIR) spectrometers operating in diffuse or trans-reflectance mode. SRM 2036 Near-Infrared Wavelength/Wavenumber Reflection Standard is a combination of a rare earth oxide glass of a composition similar to that of SRM 2035 Near-Infrared Transmission Wavelength/Wavenumber Standard and SRM 2065 Ultraviolet–Visible–Near-Infrared Transmission Wavelength/Wavenumber Standard, but is in physical contact with a piece of sintered poly(tetrafluoroethylene) (PTFE). The combination of glass contacted with a nearly ideal diffusely reflecting backing provides reflection–absorption bands that range from 15% R to 40% R. SRM 2036 is certified for the 10% band fraction air wavelength centroid location, 10%B, of seven bands spanning the spectral region from 975 nm to 1946 nm. It is also certified for the vacuum wavenumber 10%B of the same seven bands in the spectral region from 10 300 cm−1 to 5130 cm−1 at 8 cm−1 resolution. Informational values are provided for the locations of thirteen additional bands from 334 nm to 804 nm.