Fold Rise in Antibody Titers by Measured by Glycoprotein-Based Enzyme-Linked Immunosorbent Assay Is an Excellent Correlate of Protection for a Herpes Zoster Vaccine, Demonstrated via the Vaccine Efficacy Curve

The development of a micro-scale enzyme-linked immunosorbent assay (ELISA) with horseradish peroxidase as the marker enzyme for the detection and measurement of human rotavirus antibodies is described. A semipurified preparation of the serologically related simian agent, SA-11 virus, was used as the antigen. Test sera were reacted with antigen-sensitized wells in disposable poly-vinyl microplates. Any attached antibody was detected by the addition of peroxidase-labeled anti-species immunoglobulin (conjugate) followed by assay of the enzyme reaction with its substrate, hydrogen peroxide plus 5-aminosalicylic acid. This micro-ELISA was compared with complement fixation in a seroepidemiological study of the age prevalence of rotavirus antibody in Aboriginal and European populations living in the same outback area in Australia. The ELISA (results read with the naked eye) proved to be approximately 16 times more sensitive than complement fixation. Of Aborigines, 71% had rotavirus complement-fixing antibody, as compared to 45% of Europeans. By ELISA 100% of both populations had rotavirus antibodies. Mean antibody titers in the different age groups were higher in Aborigines than in Europeans. Antibody levels rose steeply throughout the first 20 years of life, remained high during the next 20 years, then increased again at least up to the age of 60 years. The micro-ELISA was practical, simple to perform, and more suitable than complement fixation for large seroepidemiological rotavirus studies. It also has potential for serodiagnosis of the disease, both in the laboratory and in the field.

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Development and Validation of 13-plex Luminex-Based Assay for Measuring Human Serum Antibodies to Streptococcus pneumoniae Capsular Polysaccharides

mSphere ◽

10.1128/msphere.00128-18 ◽

2018 ◽

Vol 3 (4) ◽

pp. e00128-18 ◽

Cited By ~ 9

Author(s):

Danka Pavliakova ◽

Peter C. Giardina ◽

Soraya Moghazeh ◽

Shite Sebastian ◽

Maya Koster ◽

...

Keyword(s):

Human Serum ◽

Lower Limit ◽

Immunosorbent Assay ◽

Enzyme Linked Immunosorbent Assay ◽

Igg Antibodies ◽

Igg Antibody ◽

Correlate Of Protection ◽

Immune Correlate ◽

Relative Standard ◽

Limit Of Quantitation

ABSTRACT A Luminex-based direct immunoassay (dLIA) platform has been developed to replace the standardized pneumococcal enzyme-linked immunosorbent assay platform. The multiplex dLIA simultaneously measures the concentration of serum immunoglobulin G (IgG) antibodies specific for pneumococcal capsular polysaccharide (PnPS) serotypes 1, 3, 4, 5, 6A, 6B, 7F, 9V, 14, 18C, 19A, 19F, and 23F. The assay uses poly-l-lysine (PLL)-conjugated PnPS, chemically coupled to spectrally distinct Luminex microspheres. Assay validation experiments were performed using residual human serum samples obtained from 13-valent pneumococcal conjugate vaccine (13vPnC) clinical studies. Assay results are expressed as IgG antibody concentrations in micrograms per milliliter using the international reference serum, 007sp. The lower limit of quantitation (LLOQ) for all serotypes covered in the 13-plex dLIA fell within the range of 0.002 to 0.038 µg/ml serum IgG. The difference between the lower limit and upper limit of the assay range was >500-fold for all serotypes, and assay variability was <20% relative standard deviation (RSD) for all serotypes. IgG antibody measurements were shown to be serotype-specific (some cross-reactivity was observed only between the structurally related serotypes 6A and 6B as well as 19A and 19F), and no interference was observed between the serotypes when the assay was performed in the 13-plex format compared to the singleplex assays. The 13-plex dLIA platform developed by Pfizer Inc. generates up to 143 test results in a single 96-well plate and is a suitable replacement of the enzyme-linked immunosorbent assay (ELISA) platform for evaluating vaccine clinical trials. IMPORTANCE The pneumococcal enzyme-linked immunosorbent assay (ELISA) measures IgG antibodies in human serum, and it is an important assay that supports licensure of pneumococcal vaccines. The immune correlate of protection, 0.35 µg/ml of IgG antibodies, was determined by the ELISA method. Pfizer has developed a new Luminex-based assay platform to replace the ELISA. These papers describe the important work of (i) validating the Luminex-based assay and (ii) bridging the immune correlate of protection (0.35 µg/ml IgG) to equivalent values reported by the Luminex platform.

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Effect of Early Immunization on Antibody Response to Reimmunization with Measles Vaccine as Demonstrated by Enzyme-Linked Immunosorbent Assay (ELISA)

PEDIATRICS ◽

10.1542/peds.74.1.90 ◽

1984 ◽

Vol 74 (1) ◽

pp. 90-93

Author(s):

M. Dianne Murphy ◽

Philip A. Brunell ◽

Alan W. Lievens ◽

Ziad M. Shehab

Keyword(s):

Antibody Response ◽

Immunosorbent Assay ◽

San Antonio ◽

Significant Risk ◽

Enzyme Linked Immunosorbent Assay ◽

Measles Vaccine ◽

Young Infants ◽

Antibody Titers

A measles epidemic in San Antonio, Texas provided a population of children who were immunized at ≤10 months of age and reimmunized at ≥15 months of age. Of these children, 302 were evaluated for measles antibody by the sensitive enzyme-linked immunosorbent assay (ELISA), and their responses were compared with those of 300 children who had been immunized at the customary time (≥15 months) with a single immunization. There were only five seronegative findings in each group. The children immunized at the customary time did have significantly higher (P < .001) antibody titers than the children immunized at ≤10 months and reimmunized at ≥15 months. These results indicate that early immunization followed by reimmunization may be indicated when young infants are at significant risk of measles exposure. This approach should not create an increased number of serologically nonresponsive children when reimmunized at ≥15 months.

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Pneumococcal Type 22F Polysaccharide Absorption Improves the Specificity of a Pneumococcal-Polysaccharide Enzyme-Linked Immunosorbent Assay

Clinical and Diagnostic Laboratory Immunology ◽

10.1128/cdli.8.2.266-272.2001 ◽

2001 ◽

Vol 8 (2) ◽

pp. 266-272 ◽

Cited By ~ 210

Author(s):

Nelydia F. Concepcion ◽

Carl E. Frasch

Keyword(s):

Correlation Coefficient ◽

Immunosorbent Assay ◽

Pneumococcal Vaccination ◽

Correlation Coefficients ◽

Enzyme Linked Immunosorbent Assay ◽

Igg Antibody ◽

Functional Activities ◽

Antibody Titers ◽

The Common ◽

Elisa Inhibition

ABSTRACT The specificity of the immune response to the 23-valent pneumococcal-polysaccharide (PS) vaccine in healthy adults and to a pneumococcal conjugate vaccine in infants was examined by measuring immunoglobulin G (IgG) antibody titers by enzyme-linked immunosorbent assay (ELISA) and the opsonophagocytosis assay. ELISA measures total antipneumococcal IgG titers including the titers of functional and nonfunctional antibodies, while the opsonophagocytosis assay measures only functional-antibody titers. Twenty-four pairs of pre- and post-pneumococcal vaccination sera from adults were evaluated (ELISA) for levels of IgG antibodies against serotypes 4, 6B, 9V, 14, 18C, 19F, and 23F. Twelve of the pairs were also examined (opsonophagocytosis assay) for their functional activities. The correlation coefficients between assay results for most types ranged from 0.75 to 0.90, but the correlation coefficient was only about 0.6 for serotypes 4 and 19F. The specificities of these antibodies were further examined by the use of competitive ELISA inhibition. A number of heterologous polysaccharides (types 11A, 12F, 15B, 22F, and 33A) were used as inhibitors. Most of the sera tested showed cross-reacting antibodies, in addition to those removed by pneumococcal C PS absorption. Our data suggest the presence of a common epitope that is found on most pneumococcal PS but that is not absorbed by purified C PS. Use of a heterologous pneumococcal PS (22F) to adsorb the antibodies to the common epitope increased the correlation between the IgG ELISA results and the opsonophagocytosis assay results. The correlation coefficient improve from 0.66 to 0.92 for type 4 and from 0.63 to 0.80 for type 19F. These common-epitope antibodies were largely absent in infants at 7 months of age, suggesting the carbohydrate nature of the epitope.

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A Modified Enzyme-Linked Immunosorbent Assay for Measurement of Antibody Responses to Meningococcal C Polysaccharide That Correlate with Bactericidal Responses

Clinical and Diagnostic Laboratory Immunology ◽

10.1128/cdli.5.4.479-485.1998 ◽

1998 ◽

Vol 5 (4) ◽

pp. 479-485 ◽

Cited By ~ 78

Author(s):

Dan M. Granoff ◽

Susan E. Maslanka ◽

George M. Carlone ◽

Brian D. Plikaytis ◽

George F. Santos ◽

...

Keyword(s):

Immunosorbent Assay ◽

Geometric Mean ◽

Correlation Coefficients ◽

Antibody Responses ◽

Enzyme Linked Immunosorbent Assay ◽

High Avidity ◽

Igg Antibody ◽

Standard Assay ◽

Bactericidal Antibody ◽

Antibody Titers

ABSTRACT The standardized enzyme-linked immunosorbent assay (ELISA) for measurement of serum immunoglobulin G (IgG) antibody responses to meningococcal C polysaccharide has been modified to employ assay conditions that ensure specificity and favor detection primarily of high-avidity antibodies. The modified and standard assays were used to measure IgG antibody concentrations in sera of toddlers vaccinated with meningococcal polysaccharide vaccine or a meningococcal C conjugate vaccine. The results were compared to the respective complement-mediated bactericidal antibody titers. In sera obtained after one or two doses of vaccine, the correlation coefficients, r, for the results of the standard assay and bactericidal antibody titers were 0.45 and 0.29, compared to 0.85 and 0.87, respectively, for the modified assay. With the standard assay, there were no significant differences between the geometric mean antibody responses of the two vaccine groups. In contrast, with the modified assay, 5- to 20-fold higher postvaccination antibody concentrations were measured in the conjugate than in the polysaccharide group. Importantly, the results of the modified assay, but not the standard ELISA, paralleled the respective geometric mean bactericidal antibody titers. Thus, by employing conditions that favor detection of higher-avidity IgG antibody, the modified ELISA provides results that correlate closely with measurements of antibody functional activity that are thought to be important in protection against meningococcal disease.

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Specific Antibodies in Sera and Gastric Aspirates of Symptomatic and Asymptomatic Helicobacter pylori-Infected Subjects

Clinical and Diagnostic Laboratory Immunology ◽

10.1128/cdli.5.3.288-293.1998 ◽

1998 ◽

Vol 5 (3) ◽

pp. 288-293 ◽

Cited By ~ 68

Author(s):

A. Mattsson ◽

A. Tinnert ◽

A. Hamlet ◽

H. Lönroth ◽

I. Bölin ◽

...

Keyword(s):

Duodenal Ulcer ◽

Helicobacter Pylori ◽

Membrane Proteins ◽

Immunosorbent Assay ◽

Antibody Responses ◽

Enzyme Linked Immunosorbent Assay ◽

Duodenal Ulcers ◽

Specific Antibodies ◽

Antibody Titers ◽

H Pylori

ABSTRACT In this study we have determined systemic and local antibody responses against different Helicobacter pylori antigens inH. pylori-infected and noninfected subjects. In addition, we studied whether differences in antibody responses between patients with duodenal ulcers and asymptomatic H. pylori carriers might explain the different outcomes of infection. Sera and in most instances gastric aspirates were collected from 19 duodenal ulcer patients, 15 asymptomatic H. pylori carriers, and 20 noninfected subjects and assayed for specific antibodies against different H. pylori antigens, i.e., whole membrane proteins (MP), lipopolysaccharides, flagellin, urease, the neuraminyllactose binding hemagglutinin HpaA, and a 26-kDa protein, by enzyme-linked immunosorbent assay. The H. pylori-infected subjects had significantly higher antibody titers against MP, flagellin, and urease in both sera and gastric aspirates compared with the noninfected subjects. Furthermore, the antibody titers against HpaA were significantly elevated in sera but not in gastric aspirates from the infected subjects. However, no differences in antibody titers against any of the tested antigens could be detected between the duodenal ulcer patients and the asymptomatic H. pyloricarriers, either in sera or in gastric aspirates.

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Development of the ELISA Test System for Quantitative Determination of IgG to the Varizella zoster virus in Human Serum and Assessment of its Diagnostic Efficiency

Epidemiology and Vaccinal Prevention ◽

10.31631/2073-3046-2021-20-6-72-80 ◽

2022 ◽

Vol 20 (6) ◽

pp. 72-80

Author(s):

L. N. Lukhverchyk ◽

G. L. Alatortseva ◽

L. N. Nesterenko ◽

V. Y. Kabargina ◽

V. V. Dotsenko ◽

...

Keyword(s):

Herpes Zoster ◽

Blood Serum ◽

Human Serum ◽

Immunosorbent Assay ◽

Test System ◽

Enzyme Linked Immunosorbent Assay ◽

Diagnostic Efficiency ◽

Functional Characteristics ◽

Serum Samples ◽

Population Immunity

Relevance. The introduction of Varicella vaccine prophylaxis explains the need to develop a methodology for monitoring the vaccination effectiveness and the intensity of population immunity. This problem can be solved using quantitative immunoassay methods. Aim. Development of an enzyme-linked immunosorbent assay for the concentration of class G immunoglobulins (AB) to Varicella zoster virus (VZV) determining and assessing its functional characteristics and diagnostic efficiency. Materials and methods. Recombinant antigen GE VZV. WHO International Standard for Antibodies to VZV W1044. Blood serum samples from healthy people and patients with Chickenpox and Herpes zoster, blood serum samples containing IgG antibodies to herpes simplex viruses of the first and second types, cytomegalovirus, Epstein-Barr virus. Anti-VZV ELISA (IgG) reagent kit (Euroimmun, Germany). Indirect enzyme-linked immunosorbent assay. Immunization of animals with recombinant antigen GE, isolation, and purification of specific antibodies. Conjugation of monoclonal antibodies to human IgG with antibodies to antigen GE and with horseradish peroxidase. Results. An enzyme-linked immunosorbent assay in «an indirect» format has been developed to determine the specific antibodies to VZV concentration (IU/ml) in human serum/plasma. An artificial calibrator for determining the concentration of AB-VZV had been synthesized and standardized according to the International WHO-standard W1044. The main functional characteristics of the developed enzyme-linked immunosorbent assay are determined in accordance with GOST 51352-2013. The diagnostic kit was tested on blood serum samples from children with chickenpox (n = 43), adults with Herpes zoster (n = 158), healthy individuals (n = 781). The diagnostic sensitivity of the test system was 85%, the diagnostic specificity was 87% according to the ROC analysis. The absence of cross-reactivity of the test system was shown on samples with serological markers of other herpesvirus infections (n = 94). Comparative trials of the developed test system and its commercial analog, the Anti-VZV ELISA (IgG) reagent kit, did not reveal statistically significant differences between their functional characteristics. Conclusions. The developed test system for determining of the AB-VZV concentration in human serum/plasma in terms of its functional characteristics meets the GOST requirements, is characterized by high diagnostic efficiency, can be used to monitor the effectiveness of vaccine prophylaxis and strength of population immunity, as well as to assess the immune response in chickenpox and Herpes zoster.

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An enzyme-linked immunosorbent assay for platelet compatibility testing

Blood ◽

10.1182/blood.v62.4.744.744 ◽

1983 ◽

Vol 62 (4) ◽

pp. 744-749 ◽

Cited By ~ 5

Author(s):

JD Tamerius ◽

JG Curd ◽

P Tani ◽

R McMillan

Keyword(s):

Antibody Titer ◽

Immunosorbent Assay ◽

Clinical Laboratory ◽

Clinical Problem ◽

Enzyme Linked Immunosorbent Assay ◽

Microtiter Plates ◽

Volunteer Donor ◽

Antibody Titers ◽

Test Serum ◽

Selection Of

Abstract The selection of platelet donors for patients who are refractory to random donor platelets often presents a difficult clinical problem. We describe an enzyme-linked immunosorbent assay (ELISA) for evaluating alloantibodies in refractory patients. Platelets from prospective donors are immobilized on microtiter plates and, after incubation with test serum and washing, platelet-bound IgG is detected with enzyme- linked anti-human IgG. Platelets from 46 prospective donors were tested. Twenty-two were judged compatible (reciprocal of the antibody titer less than 16) and, of these, 15 were used as platelet donors; each gave a measurable platelet increment after transfusion. The magnitude of the response was roughly proportional to the assay results. Platelets from donors giving antibody titers less than 4 resulted in platelet increments at 1 hr ranging from 4,890 to 22,200 (median 12,600), while platelets from donors giving titers of 8 or 16 resulted in lesser increments (550–4548). Conversely, 5 of the 24 patients found incompatible by the assay (titer greater than 16) gave no platelet increment, and in 3 instances, the recipient developed fever and chills after the transfusion. The assay is sensitive, simple, and adaptable to the clinical laboratory. Platelets from volunteer donor panels can be plated and stored for up to 6 mo.

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