Neisseria meningitidis serogroups A, B, C, Y and W135 - Serum Bactericidal Antibody assay (SBA)
Vaccines against Neisseria meningitidis capsular polysaccharide conjugates containing serogroups A, C, Y and W135 have been reported in many parts of the world. However, serogroup B Neisseria meningitidis capsular polysaccharide is non-immunogenic and at the same time has been considered a potential autoantigen, so that the vaccines are comprised of outer membrane proteins, which are type specific.
Since the early 1960s, studies Goldschneider et al., provided evidence that protection against meningococcal disease correlate with the presence of serum bactericidal activity (Goldschneider et al., 1969. Human immunity to the meningococcus. I. The role of humoral antibodies J Exp Med . 129: 1307-1326; II Development of Natural immunity J Exp Med 129: 1327-1348; IV. Immunogenicity of group A and group C meningococcal polysaccharides in human volunteers; V. The effect of immunization with meningococcal group C polysaccharides on the carrier state J Exp Med 129: 1385-1395).
Both protein and polysaccharide vaccines have been evaluated using the test of complement-mediated death in SBA assay (Serum Bactericidal Antibody assay). This test was developed in the early 1960s with the objective of determining the presence of functional specific antibodies in Neisseria meningitidis infections. Specific functional antibodies are those which in addition to binding the bacteria, activate a biological defense system such as phagocytosis (opsonophagocytosis), or activate the complement system. When the specific antibodies bind to the bacteria changes in the structure of the antibody (Fc region) promote the activation of the complement system starting fixing C1q subunit and subsequently activate the whole cascade of subunits occurs and factors (C1q, r, s, C4, C2, C3, C5, C6, C7, C8 and C9). Once you have activated all factors sequentially, bacterial die by lysis. There are several tests to determine the bactericidal activity mediated by such antibodies, with the participation of the complement system, proposed by various authors. All share three elements: bacteria, antibodies and complement. Methods of Serum bactericidal Antibody assays (SBA) differ in the number of bacteria (CFU) used; in the buffer used during the test; growth of the strain used; the incubation time of the test; the source of complement, human or rabbit; and the starting dilution of the serum. The traditional method (SBAm: Serum bactericidal microassay colony counting Antibody Assay) is considered very laborious and inappropriate when should be studied numerous samples, which involves planting and bacterial counts. For these reasons, it have been proposed alternative methods using a system to facilitate the reading of the results, such as: using triphenyltetrazolium (TTCmSBA) to display the results by the change of color of this element rather counting bacteria; using a colorimetric method (CSBA) based on the capacity of Neisseria meningitidis to consume glucose in the presence of a pH indicator to estimate the growth of the surviving bacteria.
In the studies cited previously, defined as cutoff of protection as an antibody titer ≥1:4. Subsequently, Borrow et al. (Serological basis for serogroup C meningococcal use of conjugate vaccines in the United Kingdom: reevaluation of correlates of protection Infect Immun 2001, 69: 1568-1573), established the correlation of test results using human serum or rabbit serum as complement source, and proposed for the case of using rabbit serum a cutoff protective antibody titer ≥1:8. Other authors, proposed a more conservative approach using human serum as a complement source, the cutoff proposed protection titer ≥1: 8 (Donelly, J. et al, Qualitative and quantitative assessment of the meningococcal antigens to evaluate- potential strain average of protein based vaccines PNAS, 2010, 107: 19490-19495).
For detecting functional antibodies to capsular polysaccharide (A, C, Y, W135, ...), as the polisaccharides antigenicity is constant, the Serum Bactericidal Antibody activity poses no problem, and the test results have been correlated with protection against Neisseria meningitis serogroups A, C, Y and W135 (FDA document 2011: Use of serum bactericidal antibody as an immunological correlate for demonstrating effectiveness of meningococcal conjugate vaccines -serogroups A, C, Y, and W135- administered to children less as 2 years of age). These criteria have been expanded to evaluate the protection induced by non-polysaccharide vaccines (proteins), such as vaccines against serogroup B, supporting several studies. However, a study by Perkins et al. (Immunogenicity of two efficacious outer membrane protein based serogroup B meningococcal vaccines among young adults in Iceland. J Infect Dis 1998, 177: 683-691) it suggests that serum bactericidal test the (SBA) may underestimate the clinical efficacy of the vaccine serogroup B. this may be because in the detection of functional antibodies to proteins from serogroup B, as there are several types of these proteins, could be the case that the strain used in the tests did not contain the most suitable proteins to detect antibodies present in the samples analyzed. They have attempted to evaluate several independent antigenic proteins to try to select a protein that corresponded to the most important antigen, which serum bactericidal test would provide more reliable results.
In the method of serum bactericidal activity (SBA: Serum bactericidal Antibody assay), the titer of functional antibody determined in the presence of human or rabbit complement, using serum dilutions that are faced to a suspension of Neisseria meningitidis serogroups A, C and B (or other) (capsulated), and after an incubation period, complement is added, turning to incubate and detecting the death of the bacteria, by seeding and subsequent counting of the surviving bacteria.
Tests in IVAMI
- Bactericidal antibodies (SBA: Serum Bactericidal Antibody assay) with a colony counting method, using serum from young rabbit.
Type of sample
- Serum or immunoglobulins product (2 mL).
Sample preservation
- Refrigerated: 24 to 48 hours.
- Frozen: more than 48 hours.
Delivery term
- 14 to 15 working days for a mixum of 3 to 4 samples at each time.
Cost of the test
- Consult ivami@ivami.com .