FAQ


What is the scale of the tuberculosis (TB) problem?
How is TB spread?
What is TB infection (“LTBI” or “latent TB”)?
What is TB disease (“active TB”)?
Are certain individuals at an increased risk of progressing from latent TB infection to TB disease?
Are certain groups of individuals at an increased risk of exposure to Mycobacterium tuberculosis?
How important is treatment for TB disease?
Why is the treatment period for TB disease so long?
How important is treatment for TB infection (“LTBI” or “latent TB”)?
Is there a test for the detection of TB infection (“LTBI” or “latent TB”)?
Is there a test for the detection of TB disease (“active TB”)?
What is the intended use of the T-SPOT.TB test?
What regulatory approvals does the T-SPOT.TB test have?
How quickly are T-SPOT.TB test results available?
Which blood collection tubes can be used?
Can blood collection tubes containing EDTA (purple top tubes) be used?
How much blood is needed for the T-SPOT.TB test?
What are the limitations of a tuberculin skin test (TST)?
What are the advantages of the T-SPOT.TB test over a tuberculin skin test (TST)?
Can the T-SPOT.TB test be used in place of a tuberculin skin test (TST)?
What are the advantages of the T-SPOT.TB test?
Why does the T-SPOT.TB test measure interferon-gamma?
What data is there to support the T-SPOT.TB test in clinical use?
How soon after exposure toMycobacterium tuberculosis can an infection be detected with the T-SPOT. TB test?
What action should be taken if the T-SPOT.TB test is positive?
Is a positive T-SPOT.TB test result expected in patients with a previous history of tuberculosis?
Can the T-SPOT.TB test detect infections of drug resistant tuberculosis strains such as MDR-TB, XDR-TB or TDR-TB?
Does the T-SPOT.TB test differentiate between latent TB infection and TB disease?
Are infections with mycobacteria other than Mycobacterium tuberculosis expected to produce positive T-SPOT.TB test results?
How are T-SPOT.TB test results reported?
How are T-SPOT.TB test results interpreted?
Why does the T-SPOT.TB test have a borderline for interpretation of results?
Are borderline T-SPOT.TB test results the same as invalid results?
What should I do with a borderline T-SPOT.TB test result?
What should I do with an invalid T-SPOT.TB test result?
Why does the T-SPOT.TB test include a Positive Control with each patient sample?
Why does the T-SPOT.TB test include a Nil (Negative) Control with each patient sample?
What is the sensitivity and the specificity of the T-SPOT.TB test?
Why is sensitivity important in a test for Mycobacterium tuberculosis infection?
Why is specificity important in a test for Mycobacterium tuberculosis infection?
Which criteria must be strictly applied to properly determine the specificity of interferon-gamma release assays (IGRAs)?
Are any patient groups excluded from testing with the T-SPOT.TB test?
Can the T-SPOT.TB test be used in testing samples from patients with weakened immune systems?
What is the purpose of cell washing and counting steps in the T-SPOT.TB test?
What is BCG (Bacille Calmette GuÈrin) vaccination?
Is the T-SPOT.TB test affected by previous BCG vaccination?
What is the impact of TB testing on healthcare resources?
How is the T-SPOT.TB test performed?
What is the expected frequency of invalid results with the T-SPOT.TB test?
Where can samples be sent for processing using the T-SPOT.TB test?
How much does the test cost?
Is the T-SPOT.TB test covered by insurance?

What is the scale of the tuberculosis (TB) problem?

Although effective treatment has been available for over 70 years, tuberculosis (TB) remains the second leading cause of death from an infectious disease worldwide. The World Health Organization (WHO) estimates that more than one third of the worldís population is infected with Mycobacterium tuberculosis. TB continues to be a significant disease due to factors such as immigration, the emergence of drug resistant TB strains, HIV, and other conditions that weaken the immune system.

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How is TB spread?

Tuberculosis (TB) is passed from person to person through the air. Individuals with pulmonary (lung) TB can propel aerosols containing Mycobacterium tuberculosis complex organisms into the air when they cough, sneeze, sing, speak or spit. Persons who then inhale these aerosols can become infected. Factors that determine the probability of infection include the immune status of the exposed individual, infectiousness of the TB contact and the proximity, frequency and duration of exposure.

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What is TB infection (“LTBI” or “latent TB”)?

Individuals with TB infection (“LTBI” or “latent TB”) harbor dormant Mycobacterium tuberculosis complex organisms in their bodies but are not infectious and do not have symptoms of TB disease. TB infected individuals usually have a positive T-SPOT.TB test result; however, assessing the probability of infection requires a combination of epidemiological, historical, medical and diagnostic findings. It is estimated that 10% of immunocompetent persons with latent TB infection will develop TB disease during the course of their lives. Approximately half of these individuals will develop TB disease within the first two years after infection, while the other half are at risk of developing TB disease at some stage in their life. The risk of progressing from TB infection to TB disease is increased in those with a weakened immune system. For example, patients living with HIV/AIDS are at a greatly increased risk of progressing to TB disease, as are patients who have undergone an organ transplant, or are receiving immunosuppressive therapy. When appropriately diagnosed, latent TB infection can be treated with antibiotics. Urgency of treatment is dictated by degree of risk of progression which is derived from consideration of epidemiological, medical and diagnostic findings.

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What is TB disease (“active TB”)?

Tuberculosis disease, or active TB, develops when the immune system cannot prevent Mycobacterium tuberculosis complex organisms from multiplying in the body. After exposure, persons can develop latent TB infection or TB disease. TB disease most commonly occurs in the lungs (pulmonary TB) but may occur in other body organs or spaces, particularly in immunocompromised patients or children, and may be localized or disseminated as occurs in miliary TB. Symptoms of pulmonary TB disease may include fever, cough, night sweats, weight loss, and fatigue. Without treatment, TB mortality rates are high. Individuals with TB disease usually have a positive T-SPOT.TB test result; however, assessing the probability of disease requires a combination of epidemiological, historical, medical and diagnostic findings. The definitive diagnosis of TB disease is made on the basis of isolation and identification of the TB mycobacterium in culture. Identification of the mycobacterium by other means, such as genetic tests of its presence, are increasingly being accepted as proof of infection. Persons with latent TB infection are also at risk for progression to TB disease, with that risk being modulated by age, concomitant illness, medication and other epidemiological factors.

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Are certain individuals at an increased risk of progressing from latent TB infection to TB disease?

The risk of progression to tuberculosis (TB) disease is higher in certain individuals, including:

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Are certain groups of individuals at an increased risk of exposure to Mycobacterium tuberculosis?

Yes, certain groups are more likely to be exposed to Mycobacterium tuberculosis, which may lead to tuberculosis (TB) infection and/or disease. These include:

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How important is treatment for TB disease?

Treatment for tuberculosis (TB) disease is vital. The goals of treatment include not only curing the patient, but reducing transmission to others. In general, the duration of treatment is 6 to 9 months, but is much longer in those with drug resistant TB. It is crucial that individuals with TB complete their entire course of treatment even if their symptoms improve. TB that is not adequately treated can reactivate or become resistant to drugs, making it more difficult to treat.

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Why is the treatment period for TB disease so long?

Most antibiotics capable of destroying bacteria can only do so while the bacteria are actively replicating. The replication cycle of Mycobacterium tuberculosis complex organisms is relatively long; therefore, lengthy treatment is required to ensure that all of the bacteria are destroyed. If the treatment is inconsistent or too short, some bacteria may survive, potentially allowing tuberculosis (TB) disease to reactivate or develop drug resistance.

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How important is treatment for TB infection (“LTBI” or “latent TB”)?

Treatment for latent TB infection is fundamental in preventing TB disease and overall TB elimination efforts. Completed treatment regimens reduce the risk of TB disease by up to 90%. Urgency of treatment is dictated by degree of risk of progression which is derived from consideration of epidemiological, medical and diagnostic findings.

In 2015, recognizing latent TB infection testing and treatment as critical components of ultimately eliminating TB disease, the World Health Organization (WHO) set forth its first guidelines on managing latent TB infection. (http://www.who.int/tb/publications/latent tuberculosis infection/en/).

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Is there a test for the detection of TB infection (“LTBI” or “latent TB”)?

There are several methods to detect TB infection, broadly divided into tuberculin skin tests and blood tests. A tuberculin skin test (TST), which has been used to detect TB infection for over 100 years, requires an intradermal injection of a small amount of purified protein derivative (PPD) into the skin. In 48-72 hours, the resultant induration is measured. More recently, blood based tests, referred to as interferon-gamma release assays (IGRAs), have been introduced. The technology of the T-SPOT.TB test, an IGRA, is based on the release of interferon-gamma secreted by individual effector T cells (both CD4+ and CD8+) after being stimulated by TB specific antigens.

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Is there a test for the detection of TB disease (“active TB”)?

Identification of individuals with active TB disease is critical to TB control. Those suspected of having TB disease may undergo a number of tests to confirm the diagnosis (e.g. chest x-ray, sputum culture, smear microscopy, PCR). Samples from the sputum or other sites are collected and cultured to categorically confirm the diagnosis of TB and to determine susceptibility of the strain to a range of antibiotics used for treatment. The T-SPOT.TB test may be used as a diagnostic aid in suspected TB disease patients when used in conjunction with radiography and other medical and diagnostic evaluations. The T-SPOT.TB test, like a TST and the ELISA based TB blood test, detects TB infection but does not differentiate between active TB disease and latent TB infection.

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What is the intended use of the T-SPOT.TB test?

The T-SPOT.TB test is an in vitro diagnostic test for the detection of effector T cells that respond to stimulation by Mycobacterium tuberculosis antigens ESAT-6 and CFP10 by capturing interferon-gamma secreted by effector T cells and is intended for use as an aid in the diagnosis of Mycobacterium tuberculosis infection. The T-SPOT.TB test is an indirect test for Mycobacterium tuberculosis infection (including disease) and is intended for use in conjunction with risk assessment, radiography and other medical and diagnostic evaluations.

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What regulatory approvals does the T-SPOT.TB test have?

The T-SPOT.TB test has carried the CE mark, which allows it to be marketed in the EU, since 2004. Additionally, the T-SPOT.TB test received U.S. Food and Drug Administration (FDA) premarket approval in 2008, was approved in China in 2010 and in Japan in 2012. Approvals have also been obtained in many other countries, including, but not limited to, Canada, Taiwan, Russia, Singapore, Thailand, Peru, Nigeria and Mexico.

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How quickly are T-SPOT.TB test results available?

Laboratory processing of the T-SPOT.TB test can be completed in approximately 24 hours. Most laboratories typically report results within 36 – 48 hours of sample receipt. Oxford Diagnostic Laboratories in Memphis, TN reports T-SPOT.TB test results within 36 hours of receiving the blood sample.

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Which blood collection tubes can be used?

A number of blood collection tubes are compatible with the T-SPOT.TB test, including standard sodium or lithium heparin tubes. Please refer to the package insert or contact the laboratory processing your T-SPOT.TB test samples for a complete list of acceptable blood collection tubes. Oxford Diagnostic Laboratories in Memphis, TN accepts standard green top tubes (sodium heparin and lithium heparin).

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Can blood collection tubes containing EDTA (purple top tubes) be used?

No, EDTA (ethylenediaminetetraacetic acid) affects cellsí secretion of interferon-gamma due to its chelating (calcium binding) properties. Blood collection tubes that contain this anti-coagulant therefore cannot be used.

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How much blood is needed for the T-SPOT.TB test?

Typically, in immunocompetent patients, sufficient peripheral blood mononuclear cells (PBMCs) to perform the T-SPOT.TB test can be obtained with the following age dependent guidelines:

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What are the limitations of a tuberculin skin test (TST)?

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What are the advantages of the T-SPOT.TB test over a tuberculin skin test (TST)?

The T-SPOT.TB test has a number of advantages over a tuberculin skin test (TST), including:

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Can the T-SPOT.TB test be used in place of a tuberculin skin test (TST)?

The Centers for Disease Control and Prevention (CDC) state that an interferongamma release assay (IGRA), such as the TSPOT.TB test, may be used in place of a TST in most situations requiring TB testing. The CDC prefers IGRAs over TSTs in patients at least 5 years of age that are BCGvaccinated or unlikely to return for TST reading. Additionally, the CDC states that the administrative and logistic difficulties associated with TSTs make IGRAs an attractive diagnostic aid for detecting TB. Please refer to the full guidelines for complete information.

Mazurek GH, Jereb J, Vernon A, LoBue P, Goldberg S, Castro K, Committee IE, Centers for Disease C, Prevention. Updated guidelines for using Interferon-Gamma Release Assays to detect Mycobacterium tuberculosis infection – United States, 2010. MMWR Recomm Rep 2010; 59: 1-25.

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What are the advantages of the T-SPOT.TB test?

The T-SPOT.TB test is the only TB test with sensitivity and specificity exceeding 95% in pivotal clinical studies. The T-SPOT.TB test is reliable even in challenging testing populations, including BCG-vaccinated and immunocompromised persons, and relies on routine phlebotomy procedures.

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Why does the T-SPOT.TB test measure interferon-gamma?

Interferon-gamma is crucial to immune defense against intracellular pathogens such as Mycobacterium tuberculosis. Post infection, naïve T cells become sensitized to TB-specific antigens and develop into TB-specific effector T cells (both CD4+ and CD8+), which then migrate to the site of infection and secrete interferon-gamma to activate macrophages to ingest and destroy mycobacteria. TB infected patients have TB-specific effector T cells circulating in their peripheral blood, which secrete interferon-gamma in vitro when stimulated by the antigens used in the T-SPOT.TB test. The T-SPOT.TB test directly captures interferon-gamma secreted by individual TB-specific effector T cells.

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What data is there to support the T-SPOT.TB test in clinical use?

The body of scientific evidence demonstrating the performance of the T-SPOT.TB test for the detection of TB infection (latent and active) in various patient populations continues to grow. Hundreds of peer reviewed publications, including a number of meta-analyses, have been published covering a wide range of clinical and epidemiological settings including:

  • Latent TB infection (“LTBI”)
  • TB disease (including pulmonary and extra pulmonary TB)
  • HIV infected patients
  • anti-TNF-alpha therapy candidates
  • Healthcare worker screening
  • Children, adults and the elderly
  • Malnourished individuals
  • Renal patients undergoing dialysis
  • Patients with oncological disorders
  • Low prevalence countries
  • High prevalence countries
  • TB contact tracing
  • Transplant patients

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How soon after exposure to Mycobacterium tuberculosis can an infection be detected with the T-SPOT.TB test?

The time interval for conversion following exposure is not yet well defined, but is expected to occur no later than tuberculin skin test (TST) conversion (typically 2-8 weeks).

Guidelines set forth by the Centers of Disease Control and Prevention (CDC) recommend contacts of a person with TB disease who have a negative initial interferon-gamma release assay (IGRA) or TST within 8 weeks of exposure be retested 8-10 weeks after last exposure.

Mazurek GH, Jereb J, Vernon A, LoBue P, Goldberg S, Castro K, Committee IE, Centers for Disease C, Prevention. Updated guidelines for using Interferon-Gamma Release Assays to detect Mycobacterium tuberculosis infection United States, 2010. MMWR Recomm Rep 2010; 59: 1 25.

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What action should be taken if the T-SPOT.TB test is positive?

Patients testing positive with the T-SPOT.TB test likely have TB infection and should be clinically evaluated for active TB disease. Risk of one or the other can be assessed on the basis of a combination of epidemiological, historical, medical and diagnostic findings. A definitive diagnosis of active TB disease is made on isolation and identification of the mycobacterium from the patient.

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Is a positive T-SPOT.TB test result expected in patients with a previous history of tuberculosis?

Unfortunately, there is no clear answer to this question. Studies do not consistently demonstrate that persons test negative after TB treatment. Clinical cure is described by negative sputum culture, improvement of symptoms and chest x-ray changes. This term refers to both a sterilizing cure and the return to the quiescent phase (latent TB infection). Some data have shown that a large proportion of individuals remain skin or blood test positive. The persistence of effector T cells, the cells stimulated by TB specific antigens in the T-SPOT.TB test, may suggest the presence of dormant bacteria but further study is required. Depending on requirements, other diagnostic tests and medical examinations could be considered if the patient remains positive after treatment.1-4

1. Zhang S, Shao L, Mo L, et al. Evaluation of gamma interferon release assays using Mycobacterium tuberculosis antigens for diagnosis of latent and active tuberculosis in Mycobacterium bovis BCG‑vaccinated populations. Clin Vaccine Immunol. 2010 Dec;17(12):1985‑90.

2. Bosshard V, Roux‑Lombard P, Perneger T, et al. Do results of the T‑SPOT.TB interferon‑gamma release assay change after treatment of tuberculosis? Respir Med. 2009 Jan;103(1):30‑4.

3. Chee CB, KhinMar KW, Gan SH, et al. Tuberculosis treatment effect on T‑cell interferon‑gamma responses to Mycobacterium tuberculosis‑specific antigens. Eur Respir J. 2010 Aug;36(2):355‑61.

4. Walzl G, Ronacher K, Hanekom W, Scriba TJ, Zumla A. Immunological biomarkers of tuberculosis. Nat Rev Immunol 2011; 11: 343‑354.c

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Can the T-SPOT.TB test detect infections of drug resistant tuberculosis strains such as MDR-TB, XDR-TB or TDR-TB?

Yes, the T-SPOT.TB test can detect all strains of Mycobacterium tuberculosis complex organisms, including multi drug resistant tuberculosis (MDR-TB), extensively drug resistant tuberculosis (XDR-TB) and totally drug resistant tuberculosis (TDR-TB). The antigens in the T-SPOT.TB test are common to all Mycobacterium tuberculosis strains. However, the test does not predict whether the organism is sensitive to usual drug treatments. Drug susceptibility testing occurs after isolation and identification of the organism by other methods.

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Does the T-SPOT.TB test differentiate between latent TB infection and TB disease?

No, like a tuberculin skin test (TST) and the ELISA based TB blood test, the T-SPOT.TB test does not differentiate latent TB infection from TB disease.

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Are infections with mycobacteria other than Mycobacterium tuberculosis expected to produce positive T-SPOT.TB test results?

Mycobacterium tuberculosis is the causative agent of most cases of tuberculosis and thus, T-SPOT.TB test sensitivity was determined from subjects with active, culture-confirmed Mycobacterium tuberculosis infection. Individuals infected with other Mycobacterium tuberculosis complex organisms (such as M. bovis, M. africanum, M. microti, M. canetti) usually have T cells in their blood which recognize the antigens (ESAT-6 and CFP10) used in the T-SPOT.TB test and are also anticipated to produce positive T-SPOT.TB test results. ESAT-6 and CFP10 antigens are absent from most non tuberculous mycobacteria (NTM) with the exception of M. marinum, M. szulgai and M. kansasii. While it is unclear if ESAT-6 and CFP10 are present in the genome of all subspecies of M. gordonae, it is possible that this NTM may also produce a positive result. All other non-tuberculous mycobacteria, including M. avium, are not expected to cross react with the antigens used in the T-SPOT.TB test.

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How are T-SPOT.TB test results reported?

T-SPOT.TB test results are reported as positive, negative, borderline, or invalid.

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How are T-SPOT.TB test results interpreted?

T-SPOT.TB test results are qualitative and are reported as positive, borderline (equivocal) or negative, given that the test controls perform as expected. Test results are determined by firstly enumerating the spots (captured interferon-gamma from individual T cells) in each of the patient’s four test wells (Positive Control, Nil Control, Panel A, Panel B). Spots can be counted from the test wells using a magnifying glass, stereomicroscope, or a digital image captured from a microscope. Qualitative results are interpreted by subtracting the spot count in the Nil (Negative) Control from the spot count in Panels A and B. Detailed information regarding test result interpretation can be found in the T-SPOT.TB package insert.

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Why does the T-SPOT.TB test have a borderline for interpretation of results?

The vast majority of T-SPOT.TB test results are either positive or negative.

A small percentage of test results can be borderline (equivocal), where the higher of (Panel A minus Nil Control) and (Panel B minus Nil Control) is 5, 6 or 7 spots. The borderline category is intended to reduce the likelihood of false-positive or false-negative results around the cutoff point of the T-SPOT.TB test. As opposed to an indeterminate or invalid result, a borderline result is clinically interpretable and should be followed by retesting as a substantial proportion of individuals may test positive upon retesting. In a study of US healthcare workers, 23% of subjects with a borderline test result retested as positive, suggesting the borderline category is useful in maintaining test sensitivity.1

According to the Centers of Disease Control and Prevention (CDC), “incorporation of a borderline category for the T-SPOT[.TB test] as approved by FDA increases test accuracy by classifying results near the cut point (at which small variations might affect the interpretation) as neither positive or negative.”2

1.  King TC, Upfal M, Gottlieb A, et al. T-SPOT.TB Interferon-γ Release Assay Performance in Healthcare Worker Screening at Nineteen U.S. Hospitals. Am J Respir Crit Care Med. 2015;192(3):367-373. doi:10.1164/rccm.201501-0199OC.

2. Mazurek GH, Jereb J, Vernon A, LoBue P, Goldberg S, Castro K, Committee IE, Centers for Disease C, Prevention. Updated guidelines for using Interferon-Gamma Release Assays to detect Mycobacterium tuberculosis infection United States, 2010. MMWR Recomm Rep 2010; 59: 1 25.

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Are borderline T-SPOT.TB test results the same as invalid results?

No, borderline results are clinically interpretable whereas invalid results cannot be interpreted due to the failure of the test’s Positive and/or Nil (Negative) Control. In both cases, however, retesting by collecting another blood sample is recommended.

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What should I do with a borderline T-SPOT.TB test result?

Borderline results are clinically interpretable and should be followed. Retesting by collecting another sample is recommended. In a study of U.S. healthcare workers, 23% of subjects with a borderline test result retested as positive, suggesting the borderline category is useful in maintaining test sensitivity. Upon retesting, if the test result remains borderline, other diagnostic tests and/or epidemiologic information should be used to help determine the TB infection status of the patient.1

1. King TC, Upfal M, Gottlieb A, et al. T-SPOT.TB Interferon-γ Release Assay Performance in Healthcare Worker Screening at Nineteen U.S. Hospitals. Am J Respir Crit Care Med. 2015;192(3):367-373. doi:10.1164/rccm.201501-0199OC.

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What should I do with an invalid T-SPOT.TB test result?

Invalid results are not clinically interpretable and may occur if the Positive and/or Nil (Negative) Control does not perform as expected. Retesting by collecting another sample is recommended. Upon retesting, if the test result remains invalid, other diagnostic tests and/or epidemiologic information should be used to help determine the TB infection status of the patient. Invalid results are uncommon and may be related to factors such as inappropriate blood storage conditions, delay in sample transport, patient specific conditions, or laboratory error.

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Why does the T-SPOT.TB test include a Positive Control with each patient sample?

The Positive Control serves as an indicator of patient cell functionality. Patient cells are incubated with a non specific stimulator, phytohemagglutinin, in the Positive Control sample well. The Positive Control spot count is ≥ 20 in the vast majority of patient samples. A failed Positive Control is uncommon but may be related to factors such as inappropriate blood storage conditions, delay in sample transport, technical factors or patient specific conditions. A small proportion of patients may have T cells which show only a limited response to phytohemagglutinin.

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Why does the T-SPOT.TB test include a Nil (Negative) Control with each patient sample?

The Nil (Negative) Control is designed to control for non-specific T cell reactivity. Patient cells are incubated with sterile media in the Nil Control well. For the test to be considered valid, there must be < 11 spots in the Nil Control well. A failed Nil Control is uncommon but may be related to technical factors or a patient specific condition.

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What is the sensitivity and the specificity of the T-SPOT.TB test?

The sensitivity of the T-SPOT.TB test is 95.6% and the specificity of the T-SPOT.TB test is 97.1%.

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Why is sensitivity important in a test for Mycobacterium tuberculosis infection?

Sensitivity (the ability to detect TB infected individuals) is one of the key measures of diagnostic performance. A test with high sensitivity, such as the T-SPOT.TB test, will have few false-negative results. This is especially important in patients that carry a greater risk of progression from latent TB infection to TB disease, such as those with weakened immune systems.

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Why is specificity important in a test for Mycobacterium tuberculosis infection?

In addition to sensitivity, specificity (the ability to identify non-infected individuals) is another key measure of diagnostic performance. A test with high specificity, such as the T-SPOT.TB test, will have few false-positive results.

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Which criteria must be strictly applied to properly determine the specificity of interferon-gamma release assays (IGRAs)?

When determining the specificity (ability to identify non-infected individuals) of IGRAs, it is essential that control populations only include individuals with a known low risk of tuberculosis infection. Failure to strictly apply this criteria leads to inaccurate statements concerning test specificity.

Numerous peer reviewed studies have found the specificity of the T-SPOT.TB test to be greater than 98% in low risk populations in low prevalence countries.1-5 

1. Bienek DR, Chang CK. Evaluation of an interferon-gamma release assay, T-SPOT.TB, in a population with a low prevalence of tuberculosis. Int J Tuberc Lung Dis. 2009 Nov;13(11):1416-21.

2. Wang SH, Powell DA, Nagaraja HN, Morris JD, Schlesinger LS, Turner J. Evaluation of a modified interferon-gamma release assay for the diagnosis of latent tuberculosis infection in adult and paediatric populations that enables delayed processing. Scand J Infect Dis. 2010 Dec;42(11-12):845-50.

3. Talbot EA, Harland D, Wieland-Alter W, Burrer S, Adams LV. Specificity of the tuberculin skin test and the T-SPOT.TB assay among students in a low-tuberculosis incidence setting. J Am Coll Health. 2012;60(1):94-6.

4. Mancuso JD, Mazurek GH, Tribble D, et al. Discordance among commercially available diagnostics for latent tuberculosis infection. Am J Respir Crit Care Med. 2012 Feb 15;185(4):427-34.

5. King TC, Upfal M, Gottlieb A, Adamo P, Bernacki E, Kadlecek CP, Jones JG, Humphrey-Carothers F, Rielly AF, Drewry P, Murray K, DeWitt M, Matsubara J, O’Dea L, Balser J, Wrighton-Smith P. T-SPOT®.TB Interferon-Gamma Release Assay (IGRA) Performance in Healthcare Worker Screening at 19 US Hospitals. American Journal of Respiratory and Critical Care Medicine 2015:150527134833008.

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Are any patient groups excluded from testing with the T-SPOT.TB test?

The TSPOT.TB test is not contraindicated in any patient group. Guidelines set forth by the Centers of Disease Control and Prevention (CDC) indicate that an interferongamma release assay (IGRA), such as the TSPOT.TB test, or a tuberculin skin test (TST) may be used without preference in patients 5 years of age and older. A TST is preferred in children under the age of 5, due to the limited data available in this age group, while an IGRA is preferred in patients at least 5 years of age that are either BCGvaccinated or unlikely to return for reading of a TST result.1 The American Academy of Pediatrics (AAP) also prefers a TST be used in patients under 5 years of age but acknowledge that some experts will use an IGRA in children 2 to 4 years of age, especially if they have received a BCG vaccine.2

1. Mazurek GH, Jereb J, Vernon A, LoBue P, Goldberg S, Castro K, Committee IE, Centers for Disease C, Prevention. Updated guidelines for using Interferon Gamma Release Assays to detect Mycobacterium tuberculosis infection United States, 2010. MMWR Recomm Rep 2010; 59: 125.

 2. Starke JR, Committee On Infectious D. Interferongamma release assays for diagnosis of tuberculosis infection and disease in children. Pediatrics 2014; 134: e1763177.

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Can the T-SPOT.TB test be used in testing samples from patients with weakened immune systems?

Yes, the T-SPOT.TB test is well suited for use in patients with weakened immune systems.

Each sample undergoes a cell count which is used to create a normalized (standardized and known number) suspension of cells that are subsequently incubated with TB-specific antigens. Immunocompromised patients may have a reduced number of peripheral blood mononuclear cells (PBMCs) the types of white blood cells used in the T-SPOT.TB test. In these patients, multiple blood tubes can be pooled to obtain the required number of cells to perform the test.

Pivotal clinical study data submitted to the US Food and Drug Administration (FDA) for pre market approval extensively evaluated the T-SPOT.TB test in immunocompromised individuals including, but not limited to, subjects with HIV, silicosis, diabetes, end stage renal disease and organ transplant. A negative tuberculin skin test was associated with being immunocompromised; while, no association was observed between T-SPOT.TB test result and immunocompromised status.

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What is the purpose of cell the washing and counting steps in the T-SPOT.TB test?

The cell washing step enables removal of plasma and potentially interfering substances, such as endogenous interferon-gamma, tricyclic antidepressants, and nonsteroidal anti-inflammatory drugs. After washing, the peripheral blood mononuclear cells (PBMCs) are counted to allow for an adjustment in cell concentration to correct for variations in patient PBMC counts and ensure a standard number of cells are used in the test. The washing and counting steps may be of particular importance in patients with weakened immune systems. Immunosuppressed and immunocompromised patients have a higher risk of progressing from latent TB infection to TB disease, and may be prescribed a potentially interfering substance or have a PBMC count outside of normal values.

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What is BCG (Bacille Calmette Guérin) vaccination?

Bacille CalmetteGuérin (BCG) vaccine is used in many countries with a high prevalence of TB to prevent childhood tuberculous meningitis and miliary disease, but confers limited protective value in adults. The BCG vaccine is also used as an immunotherapeutic agent for individuals with bladder cancer. BCGvaccinated individuals may produce a positive tuberculin skin test (TST), even if they are not infected with Mycobacterium tuberculosis complex organisms. This is a common cause of TST inaccuracy.

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Is the T-SPOT.TB test affected by previous BCG vaccination?

Unlike a tuberculin skin test, there is no association between BCG vaccination and TSPOT.TB test results.

The bacille CalmetteGuérin (BCG) vaccine is an attenuated derivative of virulent Mycobacterium bovis, the bovine or animal form of the TB mycobacterium. The TSPOT.TB test utilizes antigens (ESAT6 and CFP10) that are located on a genomic region designated as RD1, region of differentiation 1. The RD1 region is present in all virulent M. bovis strains but is deleted from all BCG strains. Because the antigens used in the TSPOT.TB test are not present in the BCG vaccine, the TSPOT.TB test does not produce a falsepositive result due to BCG vaccination. It should be noted, however, that patients infected with virulent M. bovis are likely to produce a positive TSPOT.TB result.

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What is the impact of TB testing on healthcare resources?

Studies have shown that the incorporation of the T-SPOT.TB test in control programs will reduce the overall cost of TB control. This is largely due to the elimination of significant indirect costs associated with a tuberculin skin test (TST) to both health care systems and patients.

TST requires the person being tested have at least two office visits. Up to one third of individuals do not return to have their test read. This may result in wasted resources and potentially dangerous gaps when containing an outbreak. Indirect resource and labor costs associated with administering and reading a TST are relatively high. These are compounded by the fact that the solution used in a TST, once opened, has to be used within a short time period leading to potential wastage of unused stock. False-negative TSTs and non returners may convert to TB disease leading to morbidity and higher costs of treating disease (including onward transmission).

False-positive TST results, often due to cross reactivity with BCG vaccine or environmental non-tuberculous mycobacteria can lead to unnecessary anti-TB treatment and associated toxicity testing and clinical follow up.

Pooran A, Booth H, Miller RF, Scott G, Badri M, Huggett JF, Rook G, Zumla A, Dheda K. Different screening strategies (single or dual) for the diagnosis of suspected latent tuberculosis: a cost effectiveness analysis. BMC Pulm Med 2010; 10: 7.

WrightonSmith P, Zellweger JP. Direct costs of three models for the screening of latent tuberculosis infection. Eur Respir J 2006; 28: 4550.

Diel R, WrightonSmith P, Zellweger JP. Costeffectiveness of interferongamma release assay testing for the treatment of latent tuberculosis. Eur Respir J 2007; 30: 321332.

FosterChang SA, Manning ML, Chandler L. Tuberculosis screening of new hospital employees: compliance, clearance to work time, and cost using tuberculin skin test and interferongamma release assays. Workplace Health Saf 2014; 62: 460467.

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How is the T-SPOT.TB test performed?

The T-SPOT.TB test is a simplified and validated ELISPOT (enzyme linked immunospot) method, which has some similarities to a conventional ELISA (enzyme linked immunosorbent assay) method. A whole blood sample is collected from the patient and sent to the laboratory. The laboratory separates the peripheral blood mononuclear cells (PBMCs), washes and counts them. A standard number of PBMCs are added to four microtiter wells, where they are exposed to a Positive Control, Nil (Negative) Control and two Mycobacterium tuberculosis specific antigens (ESAT-6 and CFP10). After overnight incubation, a conjugated antibody is added. The plate is then washed and a soluble substrate is added. Finally, the plate is washed and the numbers of spots are enumerated to determine the patientís result.

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What is the expected frequency of invalid results with the T-SPOT.TB test?

Invalid results are infrequent with the T-SPOT.TB test. Oxford Diagnostic Laboratories maintain low overall invalid rates. Contact technical support for additional information.

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Where can samples be sent for processing using the T-SPOT.TB test?

Please contact the Oxford Diagnostic Laboratories Client Support Team at 18775982522.

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How much does the test cost?

Please contact the Oxford Diagnostic Laboratories Client Support Team at 18775982522 for pricing information.

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Is the TSPOT.TB test covered by insurance?

Yes, the TSPOT.TB test is covered by the vast majority of commercial and government insurers and is described by a unique CPT® code: 86481, as “Tuberculosis test, cell mediated immunity antigen response measurement; enumeration of gamma interferonproducing Tcells in cell suspension.”

Oxford Immunotec does not make any recommendations or representation of use for any particular reimbursement code for any given test/procedure. CPT is a registered trademark of the American Medical Association.

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