Long Covid

The cellular immune system, particularly T-cells, regulates the strength of the immune response by releasing cytokines to increase or suppress the response depending on the viral load. Detection of reactive T-cells (effector cells) against a pathogen indicates contact and thus an acute or past infection, regardless of whether antibodies have been formed.

Interestingly, T-cell responses against SARS-CoV can be detected over a longer period than antibody titers. The EliSpot assay (Enzyme-linked Immuno-Spot Assay) is capable of detecting T-cell responses at the single-cell level, thus characterizing the individual immune response of infected individuals. Detection of cytokine release from T-cells stimulated with specific coronavirus antigens provides a differentiated picture of the immune response to illustrate the development of the disease course and immunity following an infection.

The advantages of the SARS-CoV-2 iSpot:

• Effective, indirect evidence of a previous COVID-19 infection
• Differentiated picture of the immune response to illustrate the progression of the disease and immunity following an infection
• Modern cellular testing method to differentiate between influenza and COVID-19 infections
• Likely cellular basis immunity - Inquiry into a base immunity conferred by the cellular immune system after previous contact with globally endemic coronaviruses
• When detecting SARS-CoV-2-specific T-cell clones, it's not only proof of prior virus exposure; the finding can also indicate existing cellular immunity.

Further information about the SARS-CoV-2 iSpot:

The pandemic outbreak of SARS-CoV-2 and its effects on the global population highlight the essential need for testing procedures capable of mapping infection courses. The particular significance lies in the associated diagnostic, prognostic, and therapeutic approaches. A comprehensive diagnosis that considers all pillars of the immune system is indispensable for determining immunity to SARS-CoV-2, in order to initiate targeted measures in the event of future local outbreaks.

The infection runs clinically asymptomatic or moderately in most cases. However, in some cases, the infection can cause a severe course leading to death. In addition to the established RT-PCR for confirming an acute infection, detecting the humoral immune response to SARS-CoV-2 is part of the diagnosis. ELISA test systems are used for antibody detection. However, their significance in terms of potential immunity and individually rapidly declining antibody titers is strongly debated among experts.

Testing the cellular immune system can be another pillar for comprehensive diagnostics. While antibodies attach to pathogens such as viruses and bacteria and destroy them, T cells attach to diseased cells in the body, such as tumor cells or virus-infected cells. T cells also help to recruit other immune cells to the area.

The CoV-iSpot developed by AID detects the IFN-γ and IL-2 responses of pathogen-specific activated T cells against SARS-CoV-2 (SARS-CoV-2 peptide mix) and Coronaviridae in general (PAN-Corona peptide mix). The SARS-CoV-2 peptide mix developed by AID contains only highly specific sequences of the immunodominant epitopes of SARS-CoV-2. To differentiate from other coronaviruses, a second peptide mix with specific sequences of conserved regions within the family of Coronaviridae (PAN-Corona peptide mix) is included in the CoV-iSpot.

The detection of reactive T cells (effector cells) against a pathogen, such as SARS-CoV-2, indicates contact and consequently an acute or past infection, regardless of whether antibodies were formed. Interferon-gamma is a pro-inflammatory cytokine mainly produced by activated T cells and NK cells. The release of IFN-γ by effector cells indicates a TH1-dominant immune response. Interleukin 2 regulates the activities of T lymphocytes. It promotes the activation and expansion of T cells and the differentiation of CD8+ T cells into memory cells. In the CoV-iSpot, the sole detection of IFN-γ-producing effector cells suggests an immune response in the context of a SARS-CoV-2 infection. The detection of IL-2 or simultaneous detection of IFN-γ and IL-2 indicates the presence of memory cells as evidence of a past SARS-CoV-2 infection with potential cellular immunity.

The clinical relevance:

The acquired immune response fundamentally differs in the humoral and cellular immune responses. The cellular immune response is T-cell dependent. T cells are divided into various subpopulations (e.g., Th1 and Th2) which release different cytokines specifically based on their function. Some of these cytokines initiate the humoral immune response. Thus, T cells are at the forefront of acquired defense mechanisms. The various T-cell subpopulations constitute the central point of both the humoral and cellular immune responses.

Effector T cells represent directly activated and differentiated T cells that possess pro-inflammatory properties. They originate from naive T cells after encountering the corresponding antigen during an infection or vaccination. Following differentiation, they proliferate into pathogen-specific T-cell clones, which are found as effector and memory T cells in peripheral blood. Regulatory T cells suppress the immune response. They, like effector T cells, arise from differentiation of naive T cells. A balance between activated effector T cells and regulatory T cells is crucial for an effective and protective immune response. An imbalanced ratio can be the basis of autoimmune disease or chronic inflammatory conditions.

The EliSpot method provides insight into these regulatory processes. Its application is very versatile, allowing for the representation of both acute illnesses and chronic inflammatory processes. Additionally, therapy monitoring is possible.

Indications:

Indirect detection of a previous COVID-19 infection

Especially in asymptomatic patients or those with mild symptoms, there are often no or only low antibody titers, which frequently decline rapidly. Many individuals already clear the coronavirus through their cellular immune system before antibodies are formed. By detecting SARS-CoV-2-specific T effector or memory cells, previous COVID-19 infections can be effectively identified.

Detection of cellular immunity

If SARS-CoV-2-specific T-cell clones can be detected, this is not only evidence of a previous virus encounter, but the finding may also indicate existing cellular immunity. This is particularly the case when evidence of memory cells is shown through the detection of IL-2 or a combined detection of IL-2 and INF-γ. According to initial studies, SARS-CoV-2-specific T-cell responses remain detectable in the CoV-iSpot for a long time. Relatively short-lived antibody responses, but long-lasting T-cell immunity, resulted in SARS patients from 2003 still having SARS-specific T memory cells found in 2020.

Indication of potential baseline immunity

In the CoV-iSpot, other human pathogenic coronaviruses are additionally used to stimulate lymphocytes, aiming to clarify the question of baseline immunity through the cellular immune system following previous contact with globally endemic coronaviruses. If PAN-Corona-specific T-cell responses are found in the CoV-iSpot and these patients do not become ill or only experience mild symptoms after a SARS-CoV-2 infection, then cellular baseline immunity seems very likely. Observations also support cellular baseline immunity, as patients with reactive PAN-Corona peptide approaches often do not show SARS-CoV-2 antibodies. Virus-infected cells are eliminated by T effector cells before plasma cell stimulation triggers antibody production.

Upcoming flu season

The upcoming flu season is likely to cause the number of suspected SARS-CoV-2 cases to reach new highs. With the CoV-iSpot, we have a modern cellular testing procedure to differentiate between influenza and COVID-19 infections. Particularly, recent findings regarding potential cellular immunity against SARS-CoV-2 and presumably existing baseline immunity through cross-reactive T cells could provide additional assurance.