An improved microspectrofluorimeter with automatic digital data logging: Construction and operation

This paper describes and discusses for microscopists and spectroscopists the choice of computer equipment and the design of programs used in the Denver Universal Microspectroradiometer (DUM). This instrument is an accurate computerized photon-counting microspectrophotometer, microspectrofluorimeter and microrefractometer. The computer is used to control the operation of the system, to acquire radiometric data of various kinds, and to reduce, analyse and output the data in a readily usable form. Since the radiometer was designed to carry out many kinds of measurements in a variety of micro- and macroscopic specimens, and since different methods of microscopy or spectroscopy have to be combined in various ways fro the study of any one specimen, no single master-program could fulfill efficiently all foreseeable requirements. Therefore, the programming developed is interactive, modular, hierarchical and hybrid. Modular interactive programming makes it possible for almost any kind of main program, applicable to almost any kind of measurement, to be assembled quickly from a collection of hierarchical subroutines. Main programs are short and composed mainly of Fortran statements calling subroutines; subroutines, in turn, automatically call other subroutines over many levels. The subroutines are independently written and optimized for maximum operational efficiency in the computer system used, or for maximum ease of transfer to other systems. This approach to programming enables someone unfamiliar with computer languages to operate the radiometric system from the console of the CRT terminal. The writing of new main programs, by linking groups of existing subroutines, requires only a minimum acquaintance with Fortran; only the writing and revision of subroutines requires programming experience. Differences and similarities in the method of computer operation between the present system and other computerized radiometers are briefly discussed.

Section: Denver Universal Microspectroradiometer ( D U M ) Iimentioning

confidence: 99%

The Denver Universal Microspectroradiometer (DUM): II. Computer configuration and modular programming for radiometry

Galbraith

Geyer

David

1975

“…Corrected values for fluorescent spectra obtained in one laboratory can thus be compared with corresponding results obtained in another; the use of a calibrated instrument also facilitates comparisons between various methods of tissue preparation. The use of calibrated microspectrofluorometers has been reported by Ritztn (1967), Bjorkland, Ehinger & Falck (1968), Rost &Pearse (1971), andVan Orden (1970); this instrument has emission correction only. The instruments described so far rely for correction on the application of either digital or manual processing of data.…”

Section: S U M M a R Ymentioning

confidence: 99%

“…The output of the function generator and the crude emission data are fed after suitable amplification to an analogue multiplier. The output of this multiplier comprising the corrected emission data is recorded synchronously on the Y (energy) axis of the recorder.In setting up the instrument careful alignment of the optical system, adjustment of illumination and elimination of stray light using procedures similar to those outlined byRost & Pearse (1971) are employed. Preparations are surveyed using low intensity red light, and the area to be examined scanned rapidly under blue light before selecting fluorescent cells for spectral analysis, so that photodecomposition of fluorescent derivatives of amines is reduced as far as possible.…”

mentioning

confidence: 99%

A microspectrofluorometer with on line real time correction of spectra

Wreford

Schofield

1975

“…However, the field of fluorescence microscopy has proceeded from simple instruments that utilize filter combinations for isolation of excitation and emission wavelengths to more sophisticated grating instruments capable of quantitating small amounts of sample fluorophors. Examples of such instruments are reported by Chance & Legallais (1959), Olson (1960), Agroskin & Korolev (1961), Caspersson, Lomakka & Rigler, 1965), Runge (1966), Goldman (1967), Thieme (1966), Ritzen (1967), Bjorklund, Ettinger & Falck (1968), Pearse & Rost (1969), Van Orden (1970 and Rost & Pearse (1971). Two of these are designed for direct recording of corrected excitation spectra (Ritzin, 1967;Bjorklund et al, 1968), while a third, with the aid of an analogue computer, will present both corrected emission and excitation spectra (Rost & Pearse, 1971).…”

Section: To Introductionmentioning

confidence: 99%

A direct recording corrected microspectrofluorometer

Mayer¹,

Novacek²

1974

Summary The construction of a microspectrofluorometer with automatic correction capabilities is given. The instrument uses a quantum counting system for correction of excitation spectra and an interpolating function generator for correction of emission spectra. An inexpensive analogue computer is an integral part of the electronic portion of the instrument and allows a number of mathematical manipulations to be performed on the sample and reference signals prior to recording or display. Under the conditions stated, the sensitivity of the instrument is in the 10−9 M range for quinine sulphate irradiated at 365 nm. The linear response range for quinine sulphate is 10−3 to 10−9 M. The operational wavelength range of the machine is between 226 and 700 nm.