Validation of Increased Blood Storage Times with the T-SPOT.
            <i>TB</i>
            Assay with T-Cell
            <i>Xtend</i>
            Reagent in Individuals with Different Tuberculosis Risk Factors

Wang, Shu Hua; Stew, Samuel S.; Cyktor, Joshua C.; Carruthers, Bridget; Turner, Joanne; Restrepo, Blanca I.

doi:10.1128/jcm.00674-12

Cited by 11 publications

(13 citation statements)

References 9 publications

(13 reference statements)

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“…Results were interpreted as positive under either of the following two conditions: 1) for a nil control with 0–5 SFCs, the number of SFCs in the ESAT-6 or CFP-10 wells minus the number of SFCs in the nil control >6; or 2) for a nil control with >5 SFCs, there were more than twice as many SFCs in the ESAT-6 or CFP-10 well as in the nil control. T-SPOT.TB was run within 8 hours, and the T-cell Xtend reagent was used to extend the storage time of the sample up to 32 h and to achieve accuracy equivalent to the standard T-SPOT.TB, which was not used in our institution [ 19 – 21 ].…”

Section: Methodsmentioning

confidence: 99%

Comparison of the Sensitivity of QuantiFERON-TB Gold In-Tube and T-SPOT.TB According to Patient Age

et al. 2016

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Currently, there are two types of interferon-gamma release assays (IGRAs) in use for the detection of tuberculosis (TB) infection, the QuantiFERON-TB Gold In-Tube test (GFT-GIT) and T-SPOT.TB. Owing to contradictory reports regarding whether the results of these IGRAs are affected by the age of the patient, we aimed to determine if these two tests have age-related differences in sensitivity. We retrospectively reviewed the medical records of diagnosed TB patients who were tested using either QFT-GIT or T-SPOT.TB from February 2008 to December 2013. The positivity of the two tests was analyzed and compared with true TB infection, which was defined as active TB based on either a positive Mycobacterium culture or a positive TB polymerase chain reaction. The QFT-GIT group included 192 TB patients, and the T-SPOT.TB group included 212 TB patients. Of the patients with pulmonary TB, 76 (39.6%) were in the QFT-GIT group and 143 (67.5%) in the T-SPOT.TB group. The overall sensitivity was 80.2% for QFT-GIT and 91.0% for T.SPOT.TB. The sensitivities of QFT-GIT and T-SPOT.TB according to age group were as follows: <29 years, 93.3% and 96.7%; 30–49 years, 86.5% and 94.7%; 50–69 years, 76.8% and 87.5%; and >70 years, 68.3% and 85.7%, respectively. The trend of age-related changes in sensitivity was significant for both QFT-GIT (p = 0.004) and T.SPOT.TB (p = 0.039). However, only QFT-GIT was significantly related to age in the multivariate analysis. QFT-GIT, but not T-SPOT.TB, was significantly affected by patient age.

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Section: Methodsmentioning

confidence: 99%

Comparison of the Sensitivity of QuantiFERON-TB Gold In-Tube and T-SPOT.TB According to Patient Age

et al. 2016

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“…The manufacturer of the T-SPOT assay offers the T-Cell Xtend reagent to extend processing time up to 32 h. The T-Cell Xtend reagent is an antibody complex added to the blood sample in the laboratory to deplete "inhibitory neutrophils." Although several groups have investigated the impact of the T-Cell Xtend reagent (12)(13)(14), none has compared immediate processing to delayed processing with and without the T-Cell Xtend reagent. Therefore, the potential utility of the T-Cell Xtend reagent remains to be demonstrated.…”

Section: Sources Of Variabilitymentioning

confidence: 99%

Interferon Gamma Release Assays for Latent Tuberculosis: What Are the Sources of Variability?

2016

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Interferon gamma release assays (IGRAs) are blood-based tests intended for diagnosis of latent tuberculosis infection (LTBI).IGRAs offer logistical advantages and are supposed to offer improved specificity over the tuberculin skin test (TST). However, recent serial testing studies of low-risk individuals have revealed higher false conversion rates with IGRAs than with TST. Reproducibility studies have identified various sources of variability that contribute to nonreproducible results. Sources of variability can be broadly classified as preanalytical, analytical, postanalytical, manufacturing, and immunological. In this minireview, we summarize known sources of variability and their impact on IGRA results. We also provide recommendations on how to minimize sources of IGRA variability. Interferon gamma (IFN-␥) release assays (IGRAs) are laboratory alternatives to the tuberculin skin test (TST) for diagnosis of latent tuberculosis infection (LTBI) (1). IGRAs are ex vivo assays that measure T-cell response after overnight stimulation with antigens that are relatively specific for Mycobacterium tuberculosis. IGRAs are increasingly replacing the TST for annual screening of U.S. health care workers and are also utilized in student/employee health and public health programs and in screening of patients prior to immunosuppression (1). In addition, the IGRA conversion rate is now being used as a measure of vaccine efficacy in TB vaccine trials (2).IGRAs are supposed to offer improved specificity over the TST (1). By and large, this advantage holds true in populations that receive Mycobacterium bovis BCG vaccination after infancy (1 year of age) or receive multiple doses; the specificity of TST is compromised in such settings (1). However, in practice, IGRAs have proved less specific in low-risk North American health care workers and college students (1). Furthermore, several studies have raised concerns over high rates of IGRA reversions (1). Given the growing use of IGRAs, it is imperative that we identify the underlying sources of variability and understand their impact on IGRA accuracy.The two most widely used IGRAs include the QuantiFERON-TB Gold In-Tube (QFT-GIT) assay (Cellestis/Qiagen, Carnegie, Australia) and the T-SPOT.TB (T-SPOT) assay (Oxford Immunotec, Abingdon, United Kingdom). Although a new, four-tube version of QFT (called QFT-Plus) has been launched by Qiagen, this version is currently not available in the United States. The QFT-GIT assay, the FDA-approved version, is an enzyme-linked immunosorbent assay (ELISA)-based, whole-blood test that uses peptides from the two RD1 antigens (ESAT-6 and CFP-10) as well as peptides from one additional antigen (TB7.7 [Rv2654c]) in an in-tube format. The QFT-GIT assay consists of three tubes: the negative-control (nil) tube that measures background IFN-␥ response, the antigen tube that measures antigen-specific response, and the positive-control (mitogen) tube that measures nonspecific T-cell response. The qualitative result (negative, positive, or indeterminate) is in...

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“…PBMCs were separated through density gradient centrifugation (Hettich Rotanta 460 RS; rotor 5624) at room temperature for 15 min at 1000 g . If isolation of PBMCs was performed between 8 and 32 h after venipuncture, then a T‐Cell Xtend (Oxford Immunotec Ltd) step was performed prior to the addition of 3 ml of RPMI medium and density gradient centrifugation, as previously described . After centrifugation, the PBMC fraction was collected and washed twice in 10 ml RPMI medium.…”

Section: Methodsmentioning

confidence: 99%

“…If isolation of PBMCs started within 8 h after venipuncture, 3 ml of fresh, prewarmed (37°C) Roswell Park Memorial Institute (RPMI) medium (Life Technologies, Invitrogen, Bleiswijk, the Netherlands) was added to 5 ml of blood and, after gently mixing, transferred into a Leucosep tube (Oxford Immunotec Ltd, Abingdon, UK). PBMCs were separated through density gradient centrifugation (Hettich Rotanta 460 RS; rotor 5624) at room temperature for 15 min at 1000 g. If isolation of PBMCs was performed between 8 and 32 h after venipuncture, then a T-Cell Xtend (Oxford Immunotec Ltd) step was performed prior to the addition of 3 ml of RPMI medium and density gradient centrifugation, as previously described [20,26,27]. After centrifugation, the PBMC fraction was collected and washed twice in 10 ml RPMI medium.…”

Section: Isolation Of Peripheral Blood Mononuclear Cellsmentioning

confidence: 99%

Prospective comparison of two enzyme-linked immunosorbent spot assays for the diagnosis of Lyme neuroborreliosis

Gorkom

Voet

Sankatsing

et al. 2020

Clinical and Experimental Immunology

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Summary Commercial cellular tests are used to diagnose Lyme borreliosis (LB), but studies on their clinical validation are lacking. This study evaluated the utility of an in‐house and a commercial enzyme‐linked immunosorbent spot (ELISpot) assay for the diagnosis of Lyme neuroborreliosis (LNB). Prospectively, peripheral blood mononuclear cells (PBMCs) were isolated from patients and controls and analysed using an in‐house Borrelia ELISpot assay and the commercial LymeSpot assay. B. burgdorferi B31 whole cell lysate and a mixture of outer surface proteins were used to stimulate the PBMCs and the numbers of interferon‐gamma‐secreting T cells were measured. Results were evaluated using receiver operating characteristic (ROC) curve analysis. Eighteen active and 12 treated LNB patients, 10 healthy individuals treated for an early (mostly cutaneous) manifestation of LB in the past and 47 untreated healthy individuals were included. Both assays showed a poor diagnostic performance with sensitivities, specificities, positive and negative predictive values ranging from 44.4–66.7%, 42.0–72.5%, 21.8–33.3% and 80.5–87.0%, respectively. The LymeSpot assay performed equally poorly when the calculation method of the manufacturer was used. Both the in‐house and the LymeSpot assay are unable to diagnose active LNB or to monitor antibiotic treatment success.

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Validation of Increased Blood Storage Times with the T-SPOT. TB Assay with T-Cell Xtend Reagent in Individuals with Different Tuberculosis Risk Factors

Cited by 11 publications

References 9 publications

Comparison of the Sensitivity of QuantiFERON-TB Gold In-Tube and T-SPOT.TB According to Patient Age

Comparison of the Sensitivity of QuantiFERON-TB Gold In-Tube and T-SPOT.TB According to Patient Age

Interferon Gamma Release Assays for Latent Tuberculosis: What Are the Sources of Variability?

Prospective comparison of two enzyme-linked immunosorbent spot assays for the diagnosis of Lyme neuroborreliosis

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