t.spot covid
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t.spot covid

HIGHLIGHTS

  • TSPOT.COVID is an ELISpot interferon gamma-release assay for SARS-CoV-2
  • TSPOT.COVID identifies a T cell response to SARS-CoV-2 spike S1 and N peptides
  • 2–8 weeks post SARS-CoV-2 diagnosis TSPOT.COVID detected 98% of infections
  • In comparison, immunoglobulin G (IgG) serology detected 83% of infections in the same period
  • Cellular immune response activated sooner and lasted longer than antibodies

Abstract

Objective

To evaluate the performance of the T-SPOT.COVID test for identifying SARS-CoV-2-responsive T-cells in participants with SARS-CoV-2 infection.

Methods

The T-SPOT.COVID test uses ELISpot interferon-gamma release assay (IGRA) methodology to measure T cell responses to SARS-CoV-2 spike S1 and nucleocapsid peptides. T-SPOT.COVID and anti-N immunoglobulin (Ig) G serology tests were performed on blood from 186 patients with nucleic acid amplification test (NAAT)-confirmed-SARS-CoV-2 infection and 100 control group participants.

Results

In the 2–8 weeks after NAAT-diagnosed SARS-CoV-2 infection, the T-SPOT.COVID test detected 98.4% (63 of 64) of infected participants, while anti-N IgG serology detected 82.8%. In the first 2 weeks after diagnosis, during adaptive immune response activation, there were less reactive T-SPOT.COVID responses (75.7%, 28 of 37 infected participants) and many less seropositive responses (32.4%). Response numbers tapered after 8 weeks; however, T-SPOT.COVID test continued to detect most participants with confirmed infection (83.6%, 56 of 67) and continued to out-perform serology (52.2%). T-SPOT.COVID response due to cross-reactive T cells was ruled out by demonstrating that, of 44 control group participants with T cells responsive to 4 human common cold coronavirus peptides, only 1 was T-SPOT.COVID reactive.

Conclusion

The T-SPOT.COVID test performed well in detecting SARS-CoV-2-sensitized T-cells over many months

Introduction

Long-term protection from infectious agents, such as the SARS-CoV-2 virus, is mediated by T cells and antibody-mediated immunity of the adaptive immune system (

Sette and Crotty, 2021

). The T-SPOT.COVID test was developed to identify the presence of SARS-CoV-2-responsive T cells.

T cells contribute to the understanding of SARS-CoV-2 infections in many ways. T cells can identify past SARS-CoV-2 infections at a time when PCR tests would be negative and antibodies levels may be waning (

Dan et al., 2021

;

Gudbjartsson et al., 2020

;

Poland et al., 2020

). T cells can provide immune memory lasting for months (

Dan et al., 2021

) and perhaps years, as suggested by the discovery of T cells to the SARS-CoV-1 coronavirus 17 years after infection (

Le Bert et al., 2020

). T cells may act independently of antibodies to control a SARS-CoV-2 infection, as shown by the recovery of COVID-19 patients who lack detectable antibodies but have SARS-CoV-2-responsive T cells (

Gallais et al., 2021

;

Sekine et al., 2020

). T cells also show reactivity to numerous SARS-CoV-2 epitopes, so have the potential to protect against many SARS-CoV-2 variants (

Grifoni et al., 2020

;

Tarke et al., 2021

). T cell-based assays can probe the longevity of an immune response following a SARS-CoV-2 infection or vaccination (

Goletti et al., 2021

;

Liu et al., 2021

;

Reynolds et al., 2021

). These various roles suggest that a T cell assay can be a key contributor to SARS-CoV-2 investigations.

The T-SPOT.COVID test, an enzyme-linked immunospot (ELISpot) assay, identifies T cells in peripheral blood that release interferon-gamma (IFN-γ) in response to stimulation with SARS-CoV-2 peptides. The T-SPOT.COVID test builds on the T-SPOT platform (Oxford Immunotec) used worldwide for tuberculosis and cytomegalovirus testing and the research version, the T-SPOT Discovery SARS-CoV-2 test (

Liu et al., 2021

;

t.spot covid
t.spot covid
Wyllie et al., 2021

). The T-SPOT.COVID ELISpot methodology is performed in many laboratories and offers a standardized comparison of T cell immunity among participants. In addition, ELISpot assays normalize the number of peripheral blood mononuclear cells (PBMCs), thus maintaining test effectiveness in participants with lymphopenia, a commonly reported condition in many COVID-19 patients (

Altmann and Boyton, 2020

) and immunosuppressed people.

The objective of this study was to evaluate the ability of the T-SPOT.COVID test to detect T cell responses in participants with or without a history of SARS-CoV-2 infection and to compare the T-SPOT.COVID test results with anti-N immunoglobulin (Ig)G serology results in the first several months after infection.

Materials and Methods

2.1 Participant recruitment

Participants for this single-center, cross-sectional study were recruited from patients who had attended the outpatient Primacare medical center in Fall River, Massachusetts, USA, between November 30, 2020, and March 24, 2021, a time of high demand for COVID-19 testing. Among other healthcare services, Primacare provided COVID-19 testing to anyone wanting or required to be tested. The New England Center for Clinical Research (NECCR) invited participants to join the study if they had received a positive SARS-CoV-2 nucleic acid amplification test (NAAT) at Primacare or if NECCR deemed them to be at low risk of SARS-CoV-2 infection. As this study was run independently from the participants’ healthcare providers, clinical data such as chest x-rays and hospitalizations records were not obtained. Informed consent and study approval were obtained from the Advarra institutional review board by NECCR at Primacare.
Confirmed-infection group: A NAAT, which detects the presence of the SARS-CoV-2 virus, was used to identify people infected with SARS-CoV-2 at the time of testing (

Rai et al., 2021

). Participants in the confirmed-infection group were recruited from asymptomatic and symptomatic patients who had had a positive SARS-CoV-2 NAAT result within the past 9 months. The date of the first positive NAAT result was considered the date of diagnosis of SARS-CoV-2 infection. Blood was drawn for Abbott SARS-CoV-2 chemiluminescent microparticle immunoassay (CMIA) anti-N IgG serology and T-SPOT.COVID tests between 0 to 249 days after diagnosis.

The analysis of responses was divided into 3 time periods: 0 to 2 weeks after diagnosis (0 to 14 days); 2+ to 8 weeks after diagnosis (15 to 56 days); and 8+ weeks after diagnosis (57+ days).
Control group: Many SARS-CoV-2 studies use frozen pre-pandemic blood for control samples; however, the T-SPOT platform requires fresh blood to ensure consistent results. Therefore fresh blood was obtained from control group participants prospectively recruited from individuals with low risk of prior SARS-CoV-2 infection. Requirements for enrollment included no current or prior signs or symptoms of COVID-19, no known contact with a confirmed SARS-CoV-2-infected individual, no prior history of a positive SARS-CoV-2 NAAT, no SARS-CoV-2 vaccination, and no prior diagnosis with SARS-CoV-1 or Middle Eastern Respiratory Syndrome (MERS). In addition, the BIOHIT HealthCare SARS-CoV-2 lateral flow anti-N IgM/IgG serology test was performed at enrollment, and the 1 person with a positive BIOHIT result was not enrolled. Blood was drawn at enrollment for testing with T-SPOT.COVID and the Abbott CMIA anti-N IgG serology test and anyone with a positive serology result was excluded from the control group.

2.2 T-SPOT.COVID test

The T-SPOT.COVID test includes over 250 SARS-CoV-2 peptides (15-mer peptides overlapping by 11 amino acids) in 2 antigen peptide pools; one pool contains peptides from the spike S1 protein, including the receptor-binding domain, and the other contains peptides from the nucleocapsid protein.
Blood samples for the T-SPOT.COVID test were processed and analyzed according to the manufacturer’s instructions. Briefly, blood samples were drawn into lithium heparin tubes which were shipped overnight to Oxford Immunotec (Abingdon, UK) in temperature-controlled shipping boxes. Next, the T-Cell Xtend reagent (Oxford Immunotec) was added to the samples, and PBMCs were isolated by density gradient centrifugation, washed, counted, and 250 000 cells/well were plated into 4 wells of a 96-well plate.

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AIRBORNE ALLERGY SCREEN KIT
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test for allergies airborne screen

Overview

During allergy skin tests, your skin is exposed to suspected allergy-causing substances (allergens) and is then observed for signs of an allergic reaction.

Along with your medical history, allergy tests may be able to confirm whether a particular substance you touch, breathe, or eat is causing symptoms.

 

Allergy skin tests are widely used to help diagnose allergic conditions, including:

  • Hay fever (allergic rhinitis)
  • Allergic asthma
  • Dermatitis (eczema)
  • Food allergies
  • Penicillin allergy
  • Bee venom allergy

Skin tests are generally safe for adults and children of all ages, including infants. In certain circumstances, though, skin tests aren’t recommended. Your doctor may advise against skin testing if you:

  • Have ever had a severe allergic reaction. You may be so sensitive to certain substances that even the tiny amounts used in skin tests could trigger a life-threatening reaction (anaphylaxis).
  • Take medications that could interfere with test results. These include antihistamines, many antidepressants and some heartburn medications. Your doctor may determine that it’s better for you to continue taking these medications than to temporarily discontinue them in preparation for a skin test.
  • Have certain skin conditions. If severe eczema or psoriasis affects large areas of skin on your arms and back — the usual testing sites — there may not be enough clear, uninvolved skin to do an effective test. Other skin conditions, such as dermatographism, can cause unreliable test results.

Blood tests (in vitro immunoglobulin E antibody tests) can be useful for those who shouldn’t or can’t undergo skin tests. Blood tests aren’t used for penicillin allergy.

In general, allergy skin tests are reliable for diagnosing allergies to airborne substances, such as pollen, pet dander and dust mites. Skin testing may help diagnose food allergies. But because food allergies can be complex, you may need additional tests or procedures.

Types of allergens

Allergens are substances that can cause an allergic reaction. There are three primary types of allergens:

  • Inhaled allergens affect the body when they come in contact with the lungs or membranes of the nostrils or throat. Pollen is the most common inhaled allergen.
  • Ingested allergens are present in certain foods, such as peanuts, soy, and seafood.
  • Contact allergens must come in contact with your skin to produce a reaction. An example of a reaction from a contact allergen is the rash and itching caused by poison ivy.

Allergy tests involve exposing you to a very small amount of a particular allergen and recording the reaction.

Insect sting allergy tests 

Why allergy testing is performed

Allergies affect more than 50 million people living in the USA, according to the American College of Allergy, Asthma, and Immunology. Inhaled allergens are by far the most common type. Seasonal allergies and hay fever, which is an allergic response to pollen, affect more than 40 million Americans.

The World Allergy Organization estimates that asthma is responsible for 250,000 deaths annually. These deaths can be avoided with proper allergy care, as asthma is considered an allergic disease process.

How allergy testing is performed

An allergy test may involve either a skin test or a blood test. You may have to go on an elimination diet if your doctor thinks you might have a food allergy.

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Skin tests

Skin tests are used to identify numerous potential allergens. This includes airborne, food-related, and contact allergens. The three types of skin tests are scratch, intradermal, and patch tests.

Your doctor will typically try a scratch test first. During this test, an allergen is placed in liquid, then that liquid is placed on a section of your skin with a special tool that lightly punctures the allergen into the skin’s surface. You’ll be closely monitored to see how your skin reacts to the foreign substance. If there’s localized redness, swelling, elevation, or itchiness of the skin over the test site, you’re allergic to that specific allergen.

If the scratch test is inconclusive, your doctor may order an intradermal skin test. This test requires injecting a tiny amount of allergen into the dermis layer of your skin. Again, your doctor will monitor your reaction.

Another form of skin test is the patch test (T.R.U.E. TESTTrusted Source). This involves using adhesive patches loaded with suspected allergens and placing these patches on your skin. The patches will remain on your body after you leave your doctor’s office. The patches are then reviewed at 48 hours after application and again at 72 to 96 hours after application.

Blood tests

If there’s a chance you’ll have a severe allergic reaction to a skin test, your doctor may call for a blood test. The blood is tested in a laboratory for the presence of antibodies that fight specific allergens. This test, called ImmunoCAP, is very successful in detecting IgE antibodies to major allergens.

Elimination diet

An elimination diet may help your doctor determine which foods are causing you to have an allergic reaction. It entails removing certain foods from your diet and later adding them back in. Your reactions will help determine which foods cause problems.

 

 

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