Test Id : PN23M

This test is the preferred test for patients previously tested for Streptococcus pneumoniae serotypes (as part of follow up testing or part of pre/post vaccine assessment).

The preferred test for patients being evaluated for possible immunodeficiency or for assessment of pneumococcal vaccination response (initial evaluation) is PNTOR / Streptococcus pneumoniae IgG Antibodies, Total, with Reflex, Serum

The preferred test for patients previously tested for total Streptococcus pneumoniae antibodies (as part of follow up testing or part of pre/post vaccine assessment) is PNTO / Streptococcus pneumoniae IgG Antibodies, Total, Serum

Collection Container/Tube:

Preferred: Serum gel

Acceptable: Red top

Submission Container/Tube: Plastic vial 

Specimen Volume: 0.5 mL

Collection Instructions: Centrifuge and aliquot serum into plastic vial

If not ordering electronically, complete, print, and send a General Request (T239) with the specimen.

0.4 mL

Assessing the IgG antibody response to active immunization with nonconjugated, 23-valent pneumococcal vaccines

Assessing the IgG antibody response to active immunization with conjugated 13-valent, 15-valent and 20-valent pneumococcal vaccines

Determining the ability of an individual to produce an antibody response to polysaccharide antigens, as part of the evaluation for humoral or combined immunodeficiencies

Streptococcus pneumoniae (S pneumoniae) is a gram-positive bacterium that causes a variety of infectious diseases in children and adults, including invasive disease (bacteriemia and meningitis) and infections of the respiratory tract (pneumonia and otitis media).(1) More than 90 serotypes of S pneumoniae have been identified, based on varying polysaccharides found in the bacterial cell wall. The serotypes responsible for disease vary with age and geographic location.

Bacterial polysaccharides induce a T-cell independent type II humoral immune response. In adults and older children, bacterial polysaccharides are effective in generating an immune response that results in production of IgG antibodies and generation of long-lived plasma cells and memory B cells.(2) S pneumoniae purified polysaccharide vaccines (PPSV) that contain a total of 23 serotypes (1, 2, 3, 4, 5, 6B, 7F, 8, 9N, 9V, 10A, 11A, 12F, 14, 15B, 17F, 18C, 19A, 19F, 20, 22F, 23F, and 33F) are available; these are referred to as PPSV23.(3) These 23 serotypes were included because, as a group, they account for approximately 90% of invasive pneumococcal infections. Antibody responses develop in approximately 75% to 85% of nonimmunocompromised adults and older children approximately 4 to 6 weeks following immunization with purified polysaccharide vaccines. A meta-analysis estimated an efficacy of 74% for prevention of invasive pneumococcal disease in adults vaccinated with PPSV23.(4) In contrast, immune responses to polysaccharide antigens in children younger than 2 years of age are generally weak.

Active immunization of children younger than 2 years requires vaccines prepared of polysaccharides conjugated to an immunogenic carrier protein (Corynebacterium diphtheria strain C7), which results in a T-cell dependent antibody response.(3) In children younger than age 6, prior to the availability of routine S pneumoniae vaccination, 7 serotypes (4, 6B, 9V, 18C, 19F, and 23F) accounted for 80% of invasive disease and up to 100% of all isolates that were found to be highly resistant to treatment with penicillin. The first pneumococcal conjugated vaccine (PCV) available for children younger than age 2 contained these 7 serotypes (PCV7). The vaccine was highly effective, with invasive disease in children younger than age 5 reduced from 99 to 21 cases per 100,000 population from 1998 to 2008.(5) In addition, it was demonstrated that after PCV7 became part of the routine vaccination schedule, only 2% of invasive disease was associated with any of the serotypes present in the vaccine. Instead, approximately 61% of the invasive disease was caused by an additional 6 serotypes (1, 3, 5, 6A, 7F, and 19A). This led to development of a 13-valent conjugated vaccine, known as PCV13. More recently, additional pneumococcal conjugate vaccines have been approved, specifically 15-valent (1, 3, 4, 5, 6A, 6B, 7F, 9V, 14, 18C, 19A, 19F, 22F, 23F, 33F) and 20-valent (1, 3, 4, 5, 6A, 6B, 7F, 8, 9V, 10A, 11A, 12F, 14, 15B, 18C, 19A, 19F, 22F, 23F, and 33F) vaccines, known as PCV15 and PCV20, respectively.

Conjugated pneumococcal vaccination is included in the routine childhood schedule, with 4 doses of PCV13 or PCV15 administered at 2, 4, 6, and 12 to 15 months.(6) For adults younger than 65 years, a single dose of PCV20 or a single dose of PCV15 followed 1 year later with a single dose of PPSV23 is recommended.(7) This same pneumococcal vaccination strategy is recommended for adults 19 to 64 years of age with immunocompromising conditions, cochlear implants, cerebrospinal fluid leaks, or other chronic health conditions.

Patients with intrinsic defects in humoral immunity, such as common variable immunodeficiency, may have impaired antibody responses to pneumococcal vaccination.(8,9) Selective antibody deficiency is a recognized clinical entity in patients older than 2 years of age and is characterized by recurrent bacterial respiratory infections, absent or subnormal antibody response to a majority of polysaccharide antigens, and normal or increased immunoglobulin concentrations, including IgG subclasses, in the context of intact humoral response to protein antigens. In several other primary immunodeficiencies, including Wiskott-Aldrich syndrome, autoimmune lymphoproliferate syndrome, and DiGeorge syndrome, IgG subclass deficiencies may also result in impaired antibody responses to polysaccharide antigens.

Results are reported in mcg/mL

As a general guideline, nonimmunocompromised adults develop IgG antibodies approximately 4 to 6 weeks following nonconjugated vaccination.

Either of the following conditions is consistent with a normal response to Streptococcus pneumonia vaccination:

1. When comparing pre- and post-vaccination samples, antibody concentrations increased by at least 2-fold for

a. >50% of serotypes in children <6 years of age

b. >70% of serotypes for individuals >6 years of age

2. In either a pre- or post-vaccination sample, antibody concentrations >/= 1.0 mcg/mL for

a. >50% of serotypes for children <6 years of age

b. >70% of serotypes for individuals >6 years of age

The humoral immune response to Streptococcal pneumoniae vaccination is affected by multiple factors, including age, immune status, vaccination history, prior infections, and carrier status.

Protective concentrations of IgG antibodies, or those required to prevent infection from S pneumoniae, have not been defined for any serotype.

Quantitation of the IgG antibody response to pneumococcal serotypes does not provide any information on the functional capacity of the serotype-specific antibodies generated (opsonization efficiency).

IgG antibodies specific for the 23 serotypes included in PPSV23 are measured in this test; except for serotype 6A, IgG antibodies specific for all the serotypes in PCV13, PCV15, and PCV20 are measured in this test.

1. Weisberg SS. Pneumococcus. Dis Mon. 2007;53(10):495-502

2. Grabenstein JD and Manoff SB. Pneumococcal polysaccharide 23-valent vaccine: long-term persistence of circulating antibody and immunogenicity and safety after revaccination in adults. Vaccine. 2012;30(30):4435-4444

3. Musher DM, Anderson R, Feldman C. The remarkable history of pneumococcal vaccination: an ongoing challenge. Pneumonia. 2022;14(1):5

4. Moberley S, Holden J, Tatham DP, Andrews RM. Vaccines for preventing pneumococcal infection in adults. Cochrane Database Syst Rev. 2013;2013(1):CD000422

5. Paradiso PR. Advances in pneumococcal disease prevention: 13-valent pneumococcal conjugate vaccine for infants and children. Clin Infect Dis. 2011;52(10):1241-1247

6. Kobayashi M, Farrar JL, Gierke R, et al. Use of 15-valent pneumococcal conjugate vaccine and 20-valent pneumococcal conjugate vaccine among U.S. adults: updated recommendations of the Advisory Committee on Immunization Practices – United States, 2022. MMWR Morb Mortal Wkly Rep. 2022;71(4):109-117

7. Kobayashi M, Farrar JL, Gierke R, et al. Use of 15-valent pneumococcal conjugate vaccine among U.S. children: updated recommendations of the Advisory Committee on Immunization Practices – United States, 2022. MMWR Morb Mortal Wkly Rep. 2022;71(37):1174-1181

8. Bonilla RA, Khan DA, Ballas ZK, et al. Practice parameter for the diagnosis and management of primary immunodeficiency. J Allergy Clin Immunol. 2015;136(5):1186-1205

9. Orange JS, Ballow M, Stiehm ER, et al. Use and interpretation of diagnostic vaccination in primary immunodeficiency: a working group report of the Basic and Clinical Immunology Interest Section of the American Academy of Allergy, Asthma and Immunology. J Allergy Clin Immunol. 2012;130(3 Suppl):S1-S24

IgG antibodies to Streptococcus pneumoniae serotypes are measured by microsphere photometry (also known as bead-based multi-plex immunoassay) using pneumococcal polysaccharides conjugated to MagPlex microspheres.

This assay detects IgG antibodies to different serotypes of S pneumoniae. The serotype-specific antibodies bound in the first stage of the assay are detected with R-phycoerythrin-conjugated, anti-human IgG antibody in the second stage of the assay.

Pneumococcal polysaccharides are modified with 4-(4,6-dimethoxy[1,3,5]triazin-2-yl)-4-methylmorpholinium chloride (DMTMM) and conjugated to magnetic carboxylated polystyrene microparticles (hereinafter referred to as magnetic beads) that have been dyed into spectrally distinct regions to be identified by the Luminex FlexMap 3D (FM3D) instrument. A mixture of 24 different beads (23 coupled to a different pneumococcal polysaccharide and 1 that is not coupled, to serve as a check for non-specific binding) is prepared in 1% BSA in PBS.

Standards and controls are prepared from pools of patient serum that have previously detected antibodies for the 23 serotypes of S pneumoniae. A dilution series of standard, controls, and patient sera are added to a microwell plate.

An equal amount of the bead mixture is added and allowed to incubate. The plate is washed using 1% BSA in PBS on a magnetic plate washer to remove unbound material. A substrate containing a reporter molecule, R-phycoerythrin conjugated anti-human IgG antibody, is added to the plate. After incubation, the plate is read on the Luminex FM3D where the magnetic beads are differentiated and quantitated by 2 separate lasers. The red diode laser (635 nm) excites the fluorescent dyes contained within the beads, and the green laser (532 nm) excites the reporter molecule bound to the bead surface. Data reduction software is used to quantify the amount of IgG antibody in unknown samples by correlation to a standard curve using a 5-Parameter Logistic (PL) curve fit.(Unpublished Mayo method)

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This test was developed and its performance characteristics determined by Mayo Clinic in a manner consistent with CLIA requirements. It has not been cleared or approved by the US Food and Drug Administration.

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