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Clinical and Vaccine Immunology, October 2009, p. 1439-1442, Vol. 16, No. 10
1071-412X/09/$08.00+0     doi:10.1128/CVI.00456-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

Variation in T-SPOT.TB Spot Interpretation between Independent Observers from Different Laboratories{triangledown}

Willeke P. J. Franken,1 Steven Thijsen,2 Ron Wolterbeek,3 John J. M. Bouwman,2 Hanane el Bannoudi,1 Sandra V. Kik,4,5 Jaap T. van Dissel,1 and Sandra M. Arend1*

Department of Infectious Diseases, Leiden University Medical Center, Leiden, The Netherlands,1 Department of Medical Microbiology and Immunology, Diakonessenhuis Utrecht/Zeist, Utrecht and Zeist, The Netherlands,2 Department of Medical Statistics, Leiden University Medical Center, Leiden, The Netherlands,3 KNCV Tuberculosis Foundation, The Hague, The Netherlands,4 Center for Infection and Immunity Amsterdam, Academic Medical Center Amsterdam, Amsterdam, The Netherlands5

Received 4 December 2008/ Returned for modification 18 February 2009/ Accepted 20 August 2009

T-SPOT.TB is a specific assay for the diagnosis of tuberculosis. The assay needs to be performed with freshly isolated cells, and interpretation requires training. T-SPOT.TB has been used in various clinical-epidemiological settings, but so far no studies have evaluated the effect of interobserver variation in test reading. Our aim was to evaluate variation between different observers in reading T-SPOT.TB results. The study was nested within an ongoing cohort study, in which part of the T-SPOT.TB had been performed with frozen material. Culture plates were read visually by four different observers from two laboratories and by two automated readers. Of 313 T-SPOT.TB assays, 235 were performed with fresh cells and 78 were performed with frozen cells. No significant difference was found between results obtained with fresh cells and those obtained with frozen cells. The percentage of positive results varied between readers by maximally 15%; five/six raters were within a 6% difference in positive results. Analysis of the observed interrater differences showed that some individuals systematically counted more spots than others did. Because test interpretation includes subtraction of background values, this systematic variance had little influence on interindividual differences. The test result as positive or negative varied between independent raters, mainly due to samples with values around the cutoff. This warrants further study regarding determinants affecting the reading of T-SPOT.TB.

* Corresponding author. Mailing address: Department of Infectious Diseases, C5-P 39, Leiden University Medical Centre, P.O. Box 9600, 2300 RC Leiden, The Netherlands. Phone: 31 71 5264915. Fax: 31 71 5266758. E-mail: s.m.arend{at}lumc.nl

{triangledown} Published ahead of print on 26 August 2009.

Clinical and Vaccine Immunology, October 2009, p. 1439-1442, Vol. 16, No. 10
1071-412X/09/$08.00+0     doi:10.1128/CVI.00456-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.





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