Abstract
Methods currently used for immunofluorescent reagent standardization require subjective visual comparison of reagents with control materials. Reactivities of reagents in immunofluorescence test kits vary from manufacturer to manufacturer. To solve these problems, a quantitative immunofluorescence (QIF) method which uses a calibrated photometric system and incorporates reducing agents into the mounting medium to reduce fading was developed to replace the visual method of endpoint determination. A uranyl glass slide was used to calibrate the instrument's voltage measurements, permitting daily comparisons and measurement of the instrument reading fluctuations. The QIF method was initially tailored to the determination of serum antibodies to Toxoplasma gondii by measuring the fluorescence intensity of individual tagged organisms. The nonspecific fluorescence intensity resulting from the counterstain was eliminated by use of a red-suppressing filter. The dilution-correlated polar fluorescence component was removed by subtraction of the intensity for the matching negative control dilution from each sample dilution intensity. The QIF method showed a 94% correlation with the visual comparison method for 62 clinical specimens.
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Selected References
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- Araujo F. G., Barnett E. V., Gentry L. O., Remington J. S. False-positive anti-Toxoplasma fluorescent-antibody tests in patients with antinuclear antibodies. Appl Microbiol. 1971 Sep;22(3):270–275. doi: 10.1128/am.22.3.270-275.1971. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Durham T. M., Colvin H. M. Premarket evaluation of commercial toxoplasmosis indirect fluorescent-antibody reagents. J Clin Microbiol. 1978 Mar;7(3):255–260. doi: 10.1128/jcm.7.3.255-260.1978. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Haaijman J. J., Wijnants F. A. Inexpensive automation of the Leitz orthoplan microfluorometer using pneumatic components. J Immunol Methods. 1975 Jun;7(2-3):255–270. doi: 10.1016/0022-1759(75)90023-x. [DOI] [PubMed] [Google Scholar]
- Jensen R. H., Greenspan J. S., Moore D., 2nd, Talal N., Roubinian J. R. Quantitative immunofluorescence microscopy of renal glomeruli from mice exhibiting murine lupus erythematosus. J Immunol Methods. 1981;42(3):343–353. doi: 10.1016/0022-1759(81)90163-0. [DOI] [PubMed] [Google Scholar]
- Kaplan D. S., Picciolo G. L. Characterization of instrumentation and calibrators for quantitative microfluorometry for immunofluorescence tests. J Clin Microbiol. 1989 Mar;27(3):442–447. doi: 10.1128/jcm.27.3.442-447.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Kaufman G. I., Nester J. F. Automation of the fluorescent treponemal antibody-absorption test. J Histochem Cytochem. 1972 Apr;20(4):255–260. doi: 10.1177/20.4.255. [DOI] [PubMed] [Google Scholar]
- Lesko S. A., Li W., Zheng G., Callahan D., Kaplan D. S., Midden W. R., Strickland P. T. Quantitative immunofluorescence assay for cyclobutyldithymidine dimers in individual mammalian cells. Carcinogenesis. 1989 Apr;10(4):641–646. doi: 10.1093/carcin/10.4.641. [DOI] [PubMed] [Google Scholar]
- Picciolo G. L., Kaplan D. S. Reduction of fading of fluorescent reaction product for microphotometric quantitation. Adv Appl Microbiol. 1984;30:197–234. doi: 10.1016/s0065-2164(08)70056-x. [DOI] [PubMed] [Google Scholar]
- Schipper J., Tilders F. J., Wassink R. G., Boleij H. F., Ploem J. S. Microfluorimetric scanning of sympathetic nerve fibers: quantification of neuronal and extraneuronal fluorescence with the aid of histogram analysis. J Histochem Cytochem. 1980 Feb;28(2):124–132. doi: 10.1177/28.2.7354211. [DOI] [PubMed] [Google Scholar]
- Sulzer A. J., Wilson M., Hall E. C. Toxoplasma gondii: polar staining in fluorescent antibody test. Exp Parasitol. 1971 Apr;29(2):197–200. doi: 10.1016/0014-4894(71)90024-5. [DOI] [PubMed] [Google Scholar]
- Wilson M., Ware D. A., Walls K. W. Evaluation of commercial serodiagnostic kits for toxoplasmosis. J Clin Microbiol. 1987 Dec;25(12):2262–2265. doi: 10.1128/jcm.25.12.2262-2265.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
- van Renterghem L., van Nimmen L. Indirect immunofluorescence in toxoplasmosis: frequency, nature and specificity of polar staining. Zentralbl Bakteriol Orig A. 1976 Aug;235(4):559–565. [PubMed] [Google Scholar]