Measurement of antibody responses to vaccines is widely used as a diagnostic laboratory tool for the identification of immunodeficiency.
When immunodeficiency is suspected based on the clinical presentation, antibody concentrations resulting from previous vaccinations or infections should be assessed:
- tetanus (2738 S-ClteAb)
- diphtheria (1253 S-CodiAb)
- pneumococcal (6297 S-SpnAbVT or 6296 S-SpnAbNV)
- Haemophilus influenzae type b (3585 S-HibAb), if the patient is known to have been vaccinated
If protective antibody levels are not present, vaccine responses to these antigens should be evaluated.
Instructions for measuring vaccine responses
- Vaccine responses should be assessed from serum samples obtained immediately before vaccination and at least 4 weeks, but no later than 2 months, after vaccination.
- At least 4 months, preferably 6 months, should have elapsed since the last IgG therapy before reliable evaluation.
Limitations in interpretation
- If the patient has received significant doses of glucocorticoids or immunomodulatory IgG within the preceding 6–12 months, the response may be secondarily reduced.
- Following high-dose immunoglobulin therapy, responses may remain suppressed for up to 9 months after treatment cessation.
T cell–dependent and T cell–independent B cell responses
B cells are activated upon encountering an antigen (protein or polysaccharide) that binds to their surface receptor. Activated B cells differentiate into plasma cells that produce antibodies.
B cells may be activated directly or via T-cell help.
In T cell–dependent responses, T cells recognize antigen presented by antigen-presenting cells and enhance antibody production. Protein antigens such as tetanus and diphtheria induce T cell–dependent responses. Tetanus generally elicits stronger responses than diphtheria.
Pneumococcal polysaccharide antigens induce immune responses by directly stimulating B cells without T-cell involvement. These responses are short-lived, with antibody levels declining to baseline typically within 3–8 years. Booster doses do not enhance responses, as polysaccharide vaccines do not induce immunological memory. In children under two years, polysaccharide vaccines are poorly immunogenic.
The Hib vaccine is a conjugate vaccine in which the polysaccharide is linked to a carrier protein, enabling a T cell–dependent response. However, Hib response cannot be used as a surrogate marker for protein antigen responses.
B cell responses in immunodeficiency
In laboratory diagnostics, vaccine-induced B cell responses are evaluated. Tetanus and diphtheria antibodies reflect T cell–dependent responses, while pneumococcal responses reflect T cell–independent function.
In combined immunodeficiency, both response types are impaired.
In primary antibody deficiencies, antibody production or function is impaired. Responses to polysaccharide antigens are particularly deficient.
Immunological findings in antibody deficiencies
- In common variable immunodeficiency (CVID), serum concentrations of IgG as well as IgA and/or IgM are reduced. Approximately 90% of patients also have low IgE levels. The age of onset ranges from early childhood to late adulthood.
- In agammaglobulinaemia, measurable antibody levels are essentially absent, and B cells are (almost) completely absent from the circulation. Symptoms typically begin in early childhood, when maternally derived antibodies transferred عبر placenta are lost.
- In hyper-IgM syndromes, serum IgM levels are normal or elevated, whereas other immunoglobulin classes are absent. B-cell phenotyping shows either absence of class-switched memory B cells (AICDA, UNG) or absence of all CD27+ B cells (CD40, CD40L).
- If antibody levels are decreased but vaccine responses are normal, the condition is not considered a severe immunodeficiency. In such cases of non-specific hypogammaglobulinaemia, severe infections are generally uncommon, and there is no clear evidence that immunoglobulin replacement therapy prolongs survival.
- In some cases, antibody levels may be normal but vaccine responses impaired, with deficiency restricted to IgG subclasses, most commonly IgG2. This condition is referred to as specific antibody deficiency. The need for treatment and follow-up is determined based on the severity of the clinical presentation.
Clinical interpretation
Interpretation of vaccine responses in immunodeficiency diagnostics is based on expert recommendations and does not replace clinical judgement in individual patient management.
In the diagnostics of immunodeficiency, assessment of vaccine responses should always include the response to the 23‑valent pneumococcal polysaccharide vaccine, as these antibody concentrations are most commonly reduced in immunodeficiencies associated with susceptibility to infections.
Patients with an inherited inability to produce antibodies against polysaccharide antigens are prone to recurrent infections caused by encapsulated bacteria (e.g. Streptococcus pneumoniae).
Guidance on selection of pneumococcal antibody assays
Limitations in interpretation of pneumococcal polysaccharide vaccine response
The antibody response to pneumococcal polysaccharide vaccination develops within 4 weeks. Therefore, serum samples should be collected no earlier than 4 weeks after vaccination, but no later than 2 months.
Antibody concentrations following polysaccharide vaccination cannot be used to determine whether an individual is protected against pneumococcal disease. The threshold value of 0.35 µg/mL used in interpretation does not reflect protection but serves as a diagnostic criterion for evaluating the ability to produce antibodies to polysaccharide antigens.
If the patient has previously received a pneumococcal conjugate vaccine, the response to polysaccharide vaccination should be interpreted only for serotypes not included in the conjugate vaccine. These are serotypes 2, 8, 10A, 11A, 12F, 15B, 17F, 20 and 33F. The response is evaluated using assay 6296 S‑SpnAbNV.
Interpretation of pneumococcal polysaccharide vaccine response
The assay 6297 S‑SpnAbVT (serotypes 1, 4, 5, 6B, 7F, 9V, 14, 18C, 19F and 23F) is used to assess response to polysaccharide vaccine in patients who have received a pneumococcal polysaccharide vaccine but not a conjugate vaccine, as these serotypes are included in both vaccines.
- The vaccine response is considered normal if antibody concentrations exceed 0.35 µg/mL in more than half of the 10 serotypes tested (6–10).
If the patient has received both polysaccharide and conjugate vaccines, the response should be evaluated using assay 6296 S‑SpnAbNV.
- The response is considered normal if antibody concentrations exceed 0.35 µg/mL in more than half of the 9 serotypes tested (5–9).
Interpretation of borderline cases
In borderline cases, the proportion of serotypes with antibody concentrations exceeding the threshold corresponding to the 5th percentile of the reference population is also assessed.
The response is considered normal if more than half of the serotypes exceed the serotype-specific concentration corresponding to the 5th percentile.
If laboratory findings are borderline, the clinical presentation is decisive in the diagnosis of immunodeficiency.
Significance of threshold values
Based on current evidence, a threshold of 0.35 µg/mL is justified for defining a normal vaccine response in immunodeficiency diagnostics.
Similar strict thresholds have been used in British studies demonstrating a significant survival benefit of IgG replacement therapy. Higher thresholds reported in American literature are based on unpublished data.
In clinical practice, serious adverse effects of long-term IgG replacement therapy are observed particularly in patients where vaccine response interpretation has been less stringent.
THL aims to collect a comprehensive Finnish patient dataset in the coming years to refine the interpretation thresholds for assays 6297 S‑SpnAbVT and 6296 S‑SpnAbNV, possibly at the serotype-specific level.
Reference data
The reference dataset is based on antibody measurements in Finnish children (2–6 years, n=56; 7–17 years, n=33) and adults (>18 years, n=61) before and after pneumococcal polysaccharide vaccination.
The reference tables include geometric mean concentrations (GMC) of pneumococcal IgG antibodies (µg/mL). The percentile indicates the concentration below which 5% of the reference population values fall. The fold increase is calculated as the ratio of geometric mean concentrations before and after vaccination.
Reference values 6297 S‑SpnAb (in Finnish), Streptococcus pneumoniae (serotypes 1, 4, 5, 6B, 7F, 9V, 14, 18C, 19F and 23F)
Reference values 6296 S‑SpnAb (in Finnish), Streptococcus pneumoniae (serotypes 2, 8, 10A, 11A, 12F, 15B, 17F, 20 and 33F)
Tetanus toxin is a protein produced by Clostridium tetani, the bacterium that causes tetanus.
Corynebacterium diphtheriae, the causative agent of diphtheria, produces diphtheria toxin.
Vaccines contain inactivated forms of these toxins (toxoids). Antibodies induced by vaccination bind to the toxins produced by the bacteria, thereby preventing tissue damage, for example in the nervous system or myocardium.
When immunodeficiency is suspected based on the clinical presentation, antibody concentrations against tetanus and diphtheria toxoids resulting from prior vaccinations should be assessed.
In immunodeficiency diagnostics, antibody responses to tetanus and diphtheria toxoids are used to evaluate whether the individual has a normal T cell–dependent B cell response.
For evaluation of suspected immunodeficiency, antibodies are measured in serum samples collected before and after vaccination. Assays 2738 S‑ClteAb (tetanus) and 1253 S‑CodiAb (diphtheria) are used. Assessment of response to the 23‑valent pneumococcal polysaccharide vaccine should always be included.
Instructions for measurement of vaccine responses
- Collect a serum sample before vaccination and 2–4 weeks after vaccination, but no later than 2 months.
- Antibodies to tetanus, diphtheria, and pneumococcal polysaccharide can be measured from the same sample.
Interpretation of tetanus and diphtheria antibodies
Protective antibody levels are considered sufficient if concentrations measured before vaccination are:
- tetanus ≥ 0.1 IU/mL
- diphtheria ≥ 0.1 IU/mL
If protective levels are not present, antibody concentrations should also be measured after vaccination.
The vaccine response is considered normal if antibody levels increase at least threefold after vaccination.
Long-term protective levels are considered sufficient if post-vaccination concentrations are:
- tetanus ≥ 1 IU/mL
- diphtheria ≥ 1 IU/mL
An antibody concentration ≥ 0.1 IU/mL is generally considered protective against clinical disease for both tetanus and diphtheria, while ≥ 1 IU/mL indicates good long-term protection.
In a Finnish serum dataset collected in 2000–2001, antibody concentrations exceeded 0.1 IU/mL in more than 95% of adults aged 30–49 years for tetanus and in approximately 80% for diphtheria.
In older age groups, a smaller proportion had protective levels, particularly for diphtheria.
Antibody concentrations decline by approximately 10% per year after vaccination. To maintain protective levels (≥ 0.1 IU/mL) for approximately 20 years, post-vaccination concentrations should be ≥ 1 IU/mL.
Pneumococcal conjugate vaccine was introduced into the national immunisation programme in 2010. All children born after 1 June 2010 are eligible to receive Synflorix vaccine, which provides protection against ten pneumococcal serotypes. Vaccinations are administered at 3, 5 and 12 months of age.
Patients who have undergone haematopoietic stem cell transplantation are also protected against pneumococcal disease. In these patients, the Prevenar13 vaccine is used.
Synflorix and Prevenar13 may also be used in other age groups. Individuals aged over five years require only a single dose.
Sample collection considerations
The antibody response to PCV vaccination develops within 4 weeks. Therefore, serum samples should be collected no earlier than 4 weeks after vaccination, but no later than 2 months.
Interpretation of pneumococcal conjugate vaccine response
Interpretation of vaccine responses is limited by the lack of a clearly defined antibody concentration that confers protection against pneumococcal disease at the individual level. Previous infection and nasopharyngeal carriage may also result in antibodies to pneumococcal polysaccharides.
By comparing antibody concentrations before and after vaccination, it is possible to assess whether the individual has mounted an immune response to the pneumococcal serotypes included in the vaccine.
In the assessment of vaccine response, the response may be considered normal if antibody concentrations show, for example, a twofold increase for all vaccine serotypes tested or a fourfold increase for at least half of the serotypes.
The World Health Organization (WHO) has defined a threshold of 0.35 µg/mL for antibody concentrations. This value is used in vaccine licensing to assess immunogenicity at the population level and is based on a meta-analysis of three vaccine trials.
However, the WHO threshold cannot be used to determine whether an individual is protected against pneumococcal disease, as protective antibody levels vary depending on the clinical presentation, serotype and age of the individual.
DTaP‑IPV‑Hib, the so‑called pentavalent vaccine, includes a component that provides protection against Hib disease. It is administered to infants as part of the national immunisation programme at 3, 5 and 12 months of age.
In certain situations, a separate Hib vaccine may be used to complete vaccination in children under five years of age. This vaccine is also used to protect individuals who are at increased risk of Hib infections due to underlying disease or its treatment, for example patients who have undergone splenectomy.
Sample collection considerations
The antibody response to Hib vaccination develops within 4 weeks. Therefore, serum samples should be collected no earlier than 4 weeks after vaccination, but no later than 2 months.
Interpretation of Hib vaccine response
In interpretation of vaccine responses, an adequate antibody response is considered to be achieved when:
- post‑vaccination Hib antibody concentration is ≥ 1 µg/mL
This threshold is based on population studies in which antibody levels ≥ 1 µg/mL have been shown to correlate well with protection against invasive Hib disease.
However, it should be noted that antibody concentrations in an individual cannot directly determine protection against Hib disease. Individual factors such as age, sex, health status and genetic factors influence the antibody level required for protection.
Protection against mucosal diseases, such as otitis media, likely requires higher antibody concentrations than protection against invasive disease.