This article was downloaded by: [McGill University Library] On: 09 February 2015, At: 21:32 Publisher: Taylor & Francis Informa Ltd Registered in England and Wales Registered Number: 1072954 Registered office: Mortimer House, 37-41 Mortimer Street, London W1T 3JH, UK
Journal of Immunoassay Publication details, including instructions for authors and subscription information: http://www.tandfonline.com/loi/ljii19
An Enzyme-Linked Immunosorbent Assay for Quantitation of Haemophilus Influenzae Type b Polysaccharide-Specific IgG1 And IgG2 in Human and Infant Rhesus Monkey Sera a
a
Kathleen T. Dolan , Joan M. Staub , Timothy L. b
Schofield , Vincent I. Ahonkhai a
& Philip P. Vella
c d
, Ronald W. Ellis
a
a
Departments of Cellular and Molecular Biology , Merck Sharp and Dohme Research Laboratories West Point , PA, 19486 b
Biometric Research , Merck Sharp and Dohme Research Laboratories West Point , PA, 19486 c
Clinical Research Merck Sharp and Dohme Research Laboratories West Point , PA, 19486 d
R. W. Johnson Pharmaceutical Research Institute Rarriton , NJ, 08869 Published online: 23 Oct 2006.
To cite this article: Kathleen T. Dolan , Joan M. Staub , Timothy L. Schofield , Vincent I. Ahonkhai , Ronald W. Ellis & Philip P. Vella (1991) An Enzyme-Linked Immunosorbent Assay for Quantitation of Haemophilus Influenzae Type b Polysaccharide-Specific IgG1 And IgG2 in Human and Infant Rhesus Monkey Sera, Journal of Immunoassay, 12:4, 543-564, DOI: 10.1080/01971529108053279 To link to this article: http://dx.doi.org/10.1080/01971529108053279
Downloaded by [McGill University Library] at 21:32 09 February 2015
PLEASE SCROLL DOWN FOR ARTICLE Taylor & Francis makes every effort to ensure the accuracy of all the information (the “Content”) contained in the publications on our platform. However, Taylor & Francis, our agents, and our licensors make no representations or warranties whatsoever as to the accuracy, completeness, or suitability for any purpose of the Content. Any opinions and views expressed in this publication are the opinions and views of the authors, and are not the views of or endorsed by Taylor & Francis. The accuracy of the Content should not be relied upon and should be independently verified with primary sources of information. Taylor and Francis shall not be liable for any losses, actions, claims, proceedings, demands, costs, expenses, damages, and other liabilities whatsoever or howsoever caused arising directly or indirectly in connection with, in relation to or arising out of the use of the Content. This article may be used for research, teaching, and private study purposes. Any substantial or systematic reproduction, redistribution, reselling, loan, sub-licensing, systematic supply, or distribution in any form to anyone is expressly forbidden. Terms & Conditions of access and use can be found at http://www.tandfonline.com/page/terms-and-conditions
JOURNAL OF IMMUNOASSAY, 1 2 ( 4 ) , 543-564 (1991)
Downloaded by [McGill University Library] at 21:32 09 February 2015
AN ENZW-LINKED IMMUNOSORBENT ASSAY FOR QUANTITATION OF HAEMOPHILUS INFLUENZAE TYPE b POLYSACCHARIDE-SPECIFIC IgGl AND IgG2 IN HUMAN AND INFANT RHESUS MONKEY SERA
Kathleen T. Dolan', Joan M. Staub', Timothy L. Schofield2, Vincent I. Ahonkhai', Ronald W. Ellis' and Philip P. Vellal Departments of Cellular and Molecular Biology,' Biometric Research,* and Clinical Research' Merck Sharp and Dohme Research Laboratories West Point, PA 19486 ABSTRACT An enzyme-linked immunosorbent assay (ELISA) has been developed and validated to quantitate IgGl and IgG2 antibody to polyribosyl- ribitol phosphate (PRP), the capsular polysaccharide of HaemoDhilus influenzae type b (Hib). The sera of children and infant Rhesus monkeys immunized with an Hib conjugate vaccine composed of Hib PRP covalently linked to an outer membrane protein complex (OMPC) from Neisseria meninuitidis serogroup B (PedvaxHIB., PRP-OMPC, Merck, Sharp and Dohme Research Laboratories). The solid-phase antigen employed in the ELISA is a conjugate of PRP to human serum albumin. The enzyme-labeled antibody is alkaline phosphatase-conjugated mouse monoclonal (mAb) anti-human IgGl or IgG2. A human serum standard was calibrated using parallel titrations with a known antibody standard. The geometric mean titer (GMT) of the anti-PRP IgGl response to one dose of PedvaxHIB. was 3.87 pg/ml (n=82), 11.80 pg/ml ( ~ 6 2 and ) 14.57 pg/ml (n=74) in infants and children 12 to 17 months, 18 to 23 months and 24 months old, respecitvely. Infants 2 to 11 months old responded with an IgGl anti-PRP response of 7.10 pg/ml while infant monkeys responded with a GMT of 150.65 (n=9) after
'Current address:
R.W. Johnson Pharmaceutical Research Institute Rarriton, NJ 08869
Requests to: Philip P. Vella Department of Cellular and Molecular Biology Merck Sharp and Dohme Research Laboratories West Point, Pennsylvania 19486 543 Copyright 0 1991 by Marcel Dekker, Inc.
544
DOLAN ET AL.
two doses of vaccine. The anti-PRP IgG2 GMT responses in all groups were 24-months old likewise produce similar
546
DOLAN ET AL.
concentrations of anti-PRP IgGl and IgG2 (8).
However, the
primary immune response to PRP-OMPC is predominantly of the IgGl subclass in 2- to 17-month old children. This suggests that a relationship may exist between the inability to produce antibody
Downloaded by [McGill University Library] at 21:32 09 February 2015
in infants against PRP vaccines (T-independentantigens) and the elicitation of primarily anti-PRP IgG1, typical of a T-dependent immune response, by Hib conjugate vaccines (3,4,5).
In order to confirm and extend these observations, we have developed and validated an ELISA to measure anti-PRP I g G l and IgG2 in PRP-OMPC-vaccinatedchildren of four different age groups and in the preclinical infant Rhesus monkey immunogenicity model.
The
assays have been standardized and validated to permit easier comparison o f such data in the field, as was done recently by the U.S. Food and Drug Administration Center for Biologicals
Evaluation and Research (CBER) for the Farr-type radioimmunoassay (RIA) for anti-PRP. MATERIALS AND METHODS Vaccine Three clinical lots of PRP-OMPC (PedvaxHIBa Lot 1072/C-P298, 1080/C-P749, and 1085/C-R132) were used to conduct clinical studies. The vaccine was a lyophilized product and was
+++/dose)
resuspended with aluminum hydroxide (0.225 mg A1 prior to use and contained 15 pg PRP/O.5 ml. administered intramuscularly into the thigh.
diluent
The vaccine was
ELISA FOR QUANTITATION OF HAEMOPHILUS INFLUENZAE
547
Human Subiects Children aged 2 months to 5 years were vaccinated with either one dose (212 mo.) or two doses (
Journal of Immunoassay Publication details, including instructions for authors and subscription information: http://www.tandfonline.com/loi/ljii19
An Enzyme-Linked Immunosorbent Assay for Quantitation of Haemophilus Influenzae Type b Polysaccharide-Specific IgG1 And IgG2 in Human and Infant Rhesus Monkey Sera a
a
Kathleen T. Dolan , Joan M. Staub , Timothy L. b
Schofield , Vincent I. Ahonkhai a
& Philip P. Vella
c d
, Ronald W. Ellis
a
a
Departments of Cellular and Molecular Biology , Merck Sharp and Dohme Research Laboratories West Point , PA, 19486 b
Biometric Research , Merck Sharp and Dohme Research Laboratories West Point , PA, 19486 c
Clinical Research Merck Sharp and Dohme Research Laboratories West Point , PA, 19486 d
R. W. Johnson Pharmaceutical Research Institute Rarriton , NJ, 08869 Published online: 23 Oct 2006.
To cite this article: Kathleen T. Dolan , Joan M. Staub , Timothy L. Schofield , Vincent I. Ahonkhai , Ronald W. Ellis & Philip P. Vella (1991) An Enzyme-Linked Immunosorbent Assay for Quantitation of Haemophilus Influenzae Type b Polysaccharide-Specific IgG1 And IgG2 in Human and Infant Rhesus Monkey Sera, Journal of Immunoassay, 12:4, 543-564, DOI: 10.1080/01971529108053279 To link to this article: http://dx.doi.org/10.1080/01971529108053279
Downloaded by [McGill University Library] at 21:32 09 February 2015
PLEASE SCROLL DOWN FOR ARTICLE Taylor & Francis makes every effort to ensure the accuracy of all the information (the “Content”) contained in the publications on our platform. However, Taylor & Francis, our agents, and our licensors make no representations or warranties whatsoever as to the accuracy, completeness, or suitability for any purpose of the Content. Any opinions and views expressed in this publication are the opinions and views of the authors, and are not the views of or endorsed by Taylor & Francis. The accuracy of the Content should not be relied upon and should be independently verified with primary sources of information. Taylor and Francis shall not be liable for any losses, actions, claims, proceedings, demands, costs, expenses, damages, and other liabilities whatsoever or howsoever caused arising directly or indirectly in connection with, in relation to or arising out of the use of the Content. This article may be used for research, teaching, and private study purposes. Any substantial or systematic reproduction, redistribution, reselling, loan, sub-licensing, systematic supply, or distribution in any form to anyone is expressly forbidden. Terms & Conditions of access and use can be found at http://www.tandfonline.com/page/terms-and-conditions
JOURNAL OF IMMUNOASSAY, 1 2 ( 4 ) , 543-564 (1991)
Downloaded by [McGill University Library] at 21:32 09 February 2015
AN ENZW-LINKED IMMUNOSORBENT ASSAY FOR QUANTITATION OF HAEMOPHILUS INFLUENZAE TYPE b POLYSACCHARIDE-SPECIFIC IgGl AND IgG2 IN HUMAN AND INFANT RHESUS MONKEY SERA
Kathleen T. Dolan', Joan M. Staub', Timothy L. Schofield2, Vincent I. Ahonkhai', Ronald W. Ellis' and Philip P. Vellal Departments of Cellular and Molecular Biology,' Biometric Research,* and Clinical Research' Merck Sharp and Dohme Research Laboratories West Point, PA 19486 ABSTRACT An enzyme-linked immunosorbent assay (ELISA) has been developed and validated to quantitate IgGl and IgG2 antibody to polyribosyl- ribitol phosphate (PRP), the capsular polysaccharide of HaemoDhilus influenzae type b (Hib). The sera of children and infant Rhesus monkeys immunized with an Hib conjugate vaccine composed of Hib PRP covalently linked to an outer membrane protein complex (OMPC) from Neisseria meninuitidis serogroup B (PedvaxHIB., PRP-OMPC, Merck, Sharp and Dohme Research Laboratories). The solid-phase antigen employed in the ELISA is a conjugate of PRP to human serum albumin. The enzyme-labeled antibody is alkaline phosphatase-conjugated mouse monoclonal (mAb) anti-human IgGl or IgG2. A human serum standard was calibrated using parallel titrations with a known antibody standard. The geometric mean titer (GMT) of the anti-PRP IgGl response to one dose of PedvaxHIB. was 3.87 pg/ml (n=82), 11.80 pg/ml ( ~ 6 2 and ) 14.57 pg/ml (n=74) in infants and children 12 to 17 months, 18 to 23 months and 24 months old, respecitvely. Infants 2 to 11 months old responded with an IgGl anti-PRP response of 7.10 pg/ml while infant monkeys responded with a GMT of 150.65 (n=9) after
'Current address:
R.W. Johnson Pharmaceutical Research Institute Rarriton, NJ 08869
Requests to: Philip P. Vella Department of Cellular and Molecular Biology Merck Sharp and Dohme Research Laboratories West Point, Pennsylvania 19486 543 Copyright 0 1991 by Marcel Dekker, Inc.
544
DOLAN ET AL.
two doses of vaccine. The anti-PRP IgG2 GMT responses in all groups were 24-months old likewise produce similar
546
DOLAN ET AL.
concentrations of anti-PRP IgGl and IgG2 (8).
However, the
primary immune response to PRP-OMPC is predominantly of the IgGl subclass in 2- to 17-month old children. This suggests that a relationship may exist between the inability to produce antibody
Downloaded by [McGill University Library] at 21:32 09 February 2015
in infants against PRP vaccines (T-independentantigens) and the elicitation of primarily anti-PRP IgG1, typical of a T-dependent immune response, by Hib conjugate vaccines (3,4,5).
In order to confirm and extend these observations, we have developed and validated an ELISA to measure anti-PRP I g G l and IgG2 in PRP-OMPC-vaccinatedchildren of four different age groups and in the preclinical infant Rhesus monkey immunogenicity model.
The
assays have been standardized and validated to permit easier comparison o f such data in the field, as was done recently by the U.S. Food and Drug Administration Center for Biologicals
Evaluation and Research (CBER) for the Farr-type radioimmunoassay (RIA) for anti-PRP. MATERIALS AND METHODS Vaccine Three clinical lots of PRP-OMPC (PedvaxHIBa Lot 1072/C-P298, 1080/C-P749, and 1085/C-R132) were used to conduct clinical studies. The vaccine was a lyophilized product and was
+++/dose)
resuspended with aluminum hydroxide (0.225 mg A1 prior to use and contained 15 pg PRP/O.5 ml. administered intramuscularly into the thigh.
diluent
The vaccine was
ELISA FOR QUANTITATION OF HAEMOPHILUS INFLUENZAE
547
Human Subiects Children aged 2 months to 5 years were vaccinated with either one dose (212 mo.) or two doses (
