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

mSphere ◽  
2018 ◽  
Vol 3 (4) ◽  
pp. e00128-18 ◽  
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.

Development of an enzyme-linked immunosorbent assay for the detection of IgG-antibodies to the causative agent of COVID-19 in human serum (plasma)

2020 ◽  
Vol 65 (11) ◽  
pp. 683-687
Author(s):  
S. G. Mardanly ◽  
A. S. Avdonina ◽  
S. G. Mamedova
Keyword(s):  
Human Serum ◽  
Immunosorbent Assay ◽  
Causative Agent ◽  
Solid Phase ◽  
Test System ◽  
Enzyme Linked Immunosorbent Assay ◽  
Diagnostic Efficiency ◽  
Igg Antibodies ◽  
Comparative Tests ◽  
Test Kit

A new original Russian test kit for the detection of IgG-antibodies to the causative agent of COVID-19 - coronavirus SARS-CoV-2 by the method of enzyme-linked immunosorbent assay (ELISA) on a solid-phase «ELISA-SARS-CoV-2-AT-G» has been developed. In comparative tests with similar test systems «Vitrotest® SARS-CoV-2 IgG» (Vitrotest, Ukraine) and «Anti-SARS-Cov-2 ELISA (IgG)» (EUROIMMUN AG, Germany) high diagnostic efficiency of the new test system was shown.

scholarly journals Evaluation of a Validated Luminex-Based Multiplex Immunoassay for Measuring Immunoglobulin G Antibodies in Serum to Pneumococcal Capsular Polysaccharides

mSphere ◽  
2018 ◽  
Vol 3 (4) ◽  
pp. e00127-18 ◽  
Author(s):  
Charles Y. Tan ◽  
Fred W. Immermann ◽  
Shite Sebastian ◽  
Michael W. Pride ◽  
Danka Pavliakova ◽  
...  
Keyword(s):  
Conjugate Vaccine ◽  
Immunosorbent Assay ◽  
Pneumococcal Conjugate Vaccine ◽  
Population Level ◽  
Enzyme Linked Immunosorbent Assay ◽  
Igg Antibodies ◽  
Specific Serum ◽  
Immune Correlate ◽  
Protective Threshold ◽  
Established Population

ABSTRACT This article describes the results of a study designed to bridge the World Health Organization (WHO) pneumococcal enzyme-linked immunosorbent assay (ELISA) platform to the validated Luminex-based 13-plex direct immunoassay (dLIA) platform developed by Pfizer, Inc. Both assay platforms quantify serotype-specific serum IgG antibodies (in micrograms per milliliter) against an international reference standard serum. The primary goal of this study was to determine if the dLIA is a suitable replacement for the ELISA to support clinical vaccine studies that include the evaluation of immune responses to serotypes 1, 3, 4, 5, 6A, 6B, 7F, 9V, 14, 18C, 19A, 19F, and 23F. Serum samples were selected from four pivotal 13-valent pneumococcal conjugate vaccine (13vPnC; Prevnar 13) clinical trials on the basis of their serotype-specific IgG concentrations by ELISA. In these studies, subjects were immunized either with 13vPnC or with 7-valent pneumococcal conjugate vaccine (7vPnC; Prevnar). There were 1,528 of 1,574 selected samples with sufficient remaining volume for reanalysis in the dLIA. A comparison of assay results from the dLIA and ELISA platforms showed clear and robust linear quantitative relationships across all 13 serotypes. In addition, lower IgG antibody concentrations in preimmunization samples were measured in the dLIA, thus allowing better differentiation between preimmunization and low-titer postimmunization samples. Overall, the results showed that the established population-level protective threshold IgG concentration, 0.35 µg/ml of serotype-specific serum IgG antibodies, is appropriate for use for data generated using the dLIA platform developed by Pfizer, Inc., for 10 serotypes: serotypes 1, 3, 4, 6A, 7F, 9V, 14, 18C, 19F, and 23F. On the basis of the extensive bridging analyses, however, the use of dLIA cutoff values of 0.23, 0.10, and 0.12 µg/ml is recommended for serotypes 5, 6B, and 19A, respectively. This adjustment will ensure that the consistency of the established population-level protective threshold IgG concentration is maintained when switching from the ELISA to the dLIA platform. The results of this bridging study demonstrate that the 13-plex dLIA platform is a suitable replacement for the WHO reference ELISA platform. 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.

scholarly journals Detection of tetanus toxoid antibodies in human sera in New Zealand by ELISA

1987 ◽  
Vol 98 (2) ◽  
pp. 199-202 ◽  
Author(s):  
R. C. H. Lau
Keyword(s):  
New Zealand ◽  
Human Serum ◽  
Immunosorbent Assay ◽  
Tetanus Toxoid ◽  
Indirect Elisa ◽  
Enzyme Linked Immunosorbent Assay ◽  
Age Group ◽  
Igg Antibodies ◽  
Serum Samples ◽  
Human Sera

SUMMARYAn enzyme-linked immunosorbent assay (ELISA) incorporating the sensitive biotin-streptavidin system was developed to detect IgG antibodies to tetanus toxoid in human serum. Serum samples obtained from 557 normal persons aged 1–65 years from different areas in New Zealand were tested. The proportion of those immune ranged from 60–93% in males, and from 46–86% in females. In the 1–9 years age group 85% were immune. The indirect ELISA is suitable for serological surveys as it is simple to perform, economical and reproducible.

Enzyme-linked immunosorbent assay for cholesteryl ester transfer protein in human serum

1995 ◽  
Vol 240 (1) ◽  
pp. 1-9 ◽  
Author(s):  
Toshitaka Sato ◽  
Masayoshi Fukasawa ◽  
Makoto Kinoshita ◽  
Hiroyuki Arai ◽  
Takao Saeki ◽  
...  
Keyword(s):  
Cholesteryl Ester ◽  
Human Serum ◽  
Immunosorbent Assay ◽  
Cholesteryl Ester Transfer Protein ◽  
Enzyme Linked Immunosorbent Assay ◽  
Transfer Protein

scholarly journals Antibody level of New Zealand children immunized with the triple vaccine DTP (diphtheria-tetanus-pertussis)

1988 ◽  
Vol 101 (2) ◽  
pp. 405-410 ◽  
Author(s):  
R. C. H Lau
Keyword(s):  
New Zealand ◽  
Immunosorbent Assay ◽  
Antibody Level ◽  
Herd Immunity ◽  
Enzyme Linked Immunosorbent Assay ◽  
Igg Antibody ◽  
Immunization Strategy ◽  
The Mean ◽  
Antibody Levels

SUMMARYEnzyme-linked immunosorbent assay (ELISA) tests were used to measure IgG antibody levels in 2638 New Zealand children who had been immunized with the triple vaccine DTP. The percentage of children immune to diphtheria decreased with age. The percentage of children immune to tetanus varied from 67.1 to 55.0%. The percentage of children with measurable antibody to pertussis increased with age. The mean percentages of children with measurable antibody or immunity to one or more DTP components were 34.2% (with 3 components), 34.4% (2 components), and 78.1% (1 component). It appears the immunization strategy for diphtheria and tetanus is satisfactory for herd immunity in New Zealand children. However, the current pertussis strategy may not be providing adequate immunity to 5-year-olds in this country.

scholarly journals Development and Application of an UHPLC-MS/MS Method for Comparative Pharmacokinetic Study of Eight Major Bioactive Components from Yin Chen Hao Tang in Normal and Acute Liver Injured Rats

2018 ◽  
Vol 2018 ◽  
pp. 1-12 ◽  
Author(s):  
Yun Wang ◽  
Xinrui Xing ◽  
Yan Cao ◽  
Liang Zhao ◽  
Sen Sun ◽  
...  
Keyword(s):  
Lower Limit ◽  
Limit Of Detection ◽  
Injury Model ◽  
Relative Standard ◽  
Limit Of Quantitation ◽  
Liquid Chromatography Tandem Mass ◽  
Bioactive Ingredients ◽  
Aloe Emodin ◽  
Plasma Pharmacokinetics ◽  
Ultrahigh Performance Liquid Chromatography

Yin Chen Hao Tang (YCHT) is one of the most famous hepatoprotective herbal formulas in China, but its pharmacokinetic investigation in model rats has been rarely conducted. In this study, the hepatic injury model was caused by intraperitoneal injections of carbon tetrachloride (CCl4), and YCHT was orally administered to the model and normal rats. An ultrahigh performance liquid chromatography–tandem mass spectrometry (UHPLC–MS/MS) method was established to analyze the plasma pharmacokinetics of eight major bioactive ingredients from YCHT in both the normal and liver injured rats. The calibration curves presented good linearity (r > 0.9981) in the concentration range. The relative standard deviation (RSD%) of inter- and intraday precision was within 9.55%, and the accuracy (RE%) ranged from -10.72% to 2.46%. The extraction recovery, matrix effect, and stability were demonstrated to be within acceptable ranges. The lower limit of detection (LLOD) and lower limit of quantitation (LLOQ) were around 0.1 ng/mL and 0.5 ng/mL, respectively, which were much lower than those in other related researches. Results reveal that there are significant differences in the pharmacokinetics of scoparone, geniposide, rhein, aloe-emodin, physcion, and chrysophanol in hepatic injured rats as compared to those in control except for scopoletin and emodin. Our experimental results provide a meaningful reference for the clinical dosage of YCHT in treating liver disorders, and the improvement of LLOD and LLOQ can also broaden the range of our method’s application, which is very suitable for quantitating these eight compounds with low levels.

Measles antibodies and response to vaccination in children aged less than 14 months: implications for age of vaccination

2011 ◽  
Vol 140 (9) ◽  
pp. 1599-1606 ◽  
Author(s):  
E. BORRÀS ◽  
L. URBIZTONDO ◽  
J. COSTA ◽  
J. BATALLA ◽  
N. TORNER ◽  
...  
Keyword(s):  
Immunosorbent Assay ◽  
Maternal Antibodies ◽  
Passive Immunity ◽  
Enzyme Linked Immunosorbent Assay ◽  
Igg Antibodies ◽  
Blood Samples ◽  
Measles Outbreak ◽  
Measles Antibodies ◽  
Measles Vaccination

SUMMARYPassive immunity against measles decreases during the first months of life. The objective of this study was to determine titres of measles antibodies in children aged 9–14 months and their mothers before vaccination, and the children's response to vaccination. Blood samples were collected by capillary puncture before and 28 days after vaccination. Samples were obtained between February and June 2007 during an ongoing measles outbreak. Titres of specific measles IgG antibodies were determined by enzyme-linked immunosorbent assay. Seroconversion was defined as the presence of antibodies after vaccination in subjects without antibodies before vaccination. Maternal antibodies were present in 37·7% of all 69 children included and in 45·1% of children aged 9 months. Of the 51 children in whom a second sample was obtained, 31 (60·8%) were seronegative before vaccination and 61·3% seroconverted. Interference of maternal antibodies was 30%. Advancing the first dose of measles vaccination from 15 to 12 months is a correct strategy, given the increase in the time of susceptibility of infants to measles.

scholarly journals Επιπολασμός λοίμωξης από ελικοβακτηρίδιο του πυλωρού σε ασθενείς με χρόνια αποφρακτική πνευμονοπάθεια

2005 ◽  
Author(s):  
Αναστάσιος Ρούσσος
Keyword(s):  
Immunosorbent Assay ◽  
Enzyme Linked Immunosorbent Assay ◽  
Igg Antibodies

Ασθενείς με πεπτικό έλκος εμφανίζουν αυξημένο επιπολασμό Χρόνιας Αποφρακτικής Πνευμονοπάθειας (ΧΑΠ) . Το κάπνισμα το οποίο αποτελεί κοινό προδιαθετικό παράγοντα και για τα δύο αυτά νοσήματα έχει ενοχοποιηθεί για τη συσχέτιση αυτή Όμως, πρόσφατες μελέτες έδειξαν ότι η ασθενείς με χρόνια βρογχίτιδα, ίσως, παρουσιάζουν αυξημένο επιπολασμό Η. pylori λοίμωξης Είναι γνωστό ότι σε ορισμένους ασθενείς η λοίμωξη από Η. pylori ενεργοποιεί την απελευθέρωση μίας σειράς προφλεγμονωδών κυτοκινών. Υπεύθυνα για την παραγωγή αυτών των κυτοκινών θεωρούνται τα ιδιαίτερα λοιμογόνα στελέχη Η. pylori τα οποία παράγουν την πρωτεΐνη CagA. Οι προφλεγμονώδεις αυτές κυτοκίνες εμπλέκονται και στην παθογένεια της ΧΑΠ, πιθανότατα προάγοντας τη μη ειδική φλεγμονή του βρογχικού δέντρου. Σκοπός της παρούσας μελέτης ήταν η διερεύνηση του επιπολασμού της Η. pylori λοίμωξης και ιδιαίτερα των CagA θετικών λοιμογόνων στελεχών σε ασθενείς με ΧΑΠ. Επίσης, επιχειρήθηκε η συσχέτιση της οροθετικότητας για το Η. pylori και για τα CagA θετικά στελέχη του με κλινικοεργαστηριακές παραμέτρους της ΧΑΠ (σπιρομετρικός έλεγχος και βαρύτητα της νόσου) Συνολικά μελετήθηκαν 126 ασθενείς με ΧΑΠ (88 άντρες and 38 γυναίκες με ηλικία 61.3 ± 8.1 έτη) και 126 μάρτυρες, προτυποποιημένοι κατά φύλο ηλικία και κοινωνικοοικονιμικό επίπεδο. Όλα τα άτομα τα οποία συμπεριελήφθησαν στη μελέτη (ασθενείς και μάρτυρες) υποβλήθηκαν σε ενζυμική ανοσοπροσροφητική μέθοδο προσδιορισμού [enzyme-linked immunosorbent assay (elisa)] των IgG αντισωμάτων για το Η. pylori και την πρωτεΐνη CagA. Επίσης, οι ασθενείς με ΧΑΠ υποβλήθηκαν σε σπιρομετρικό έλεγχο (FEV1, FEV1/FVC) και σε συνακόλουθη σταδιοποίηση της βαρύτητας της νόσου Ο εττιπολασμός της FI pylori λοίμωξης ήταν υψηλότερος στους ασθενείς με ΧΑΠ συγκριτικά με τους μάρτυρες. [77.8% έναντι 54.7% αντίστοιχα (ρ<0.001)]. Ο επιπολασμός της λοίμωξης από CagA (+) Η pylori στελέχη ήταν υψηλότερος στους ασθενείς με ΧΑΠ συγκριτικά με τους μάρτυρες [53.9% έναντι 29.3% αντίστοιχα (ρΟ.ΟΟΙ)]. Επίσης, οι ασθενείς με ΧΑΠ είχαν σημαντικά υψηλότερη μέση συγκέντρωση τόσο anti-FI. pylori IgG (118.3±24.4 vs 61.9±12.9 U/ML, ρ<0.001) όσο και anti-CagA IgG antibodies (33.8±3.4 vs 19.0±1.5 U/ML, ρ<0.001). Οι σπιρομετρικές παράμετροι δεν διέψεραν σημαντικά μεταξύ των Fi pylori (+) και Η pylori (-) ασθενών με ΧΑΠ (FEV1:61.5±18.9%, FEV1/FVC 63.0± 4.9%, έναντι FEV1:63.5±17.9%, FEV1/FVC: 63.9± 4.6%, αντίστοιχα, ρ>0.05). Παρομοίως, οι σπιρομετρικές παράμετροι δεν διέφεραν σημαντικά μεταξύ των CagA(+) και CagA (-) ασθενών με ΧΑΠ (FEV1: 59.1 ±9.7%, FEV1/FVC 62.4± 5.1%, έναντι FEV1: 65.4±6.3%, FEV1/FVC: 64.3± 4.3%, αντίστοιχα, ρ>0.05) Τέλος, δεν αναδείχθηκε σημαντική διαφορά μεταξύ των ποσοστών αντί- FI pylori IgG θετικότητας και αντι-CagA IgG θετικότητας στα διάφορα στάδια ΧΑΠ. Με δεδομένο τον υψηλό επιπολασμό της ελικοβακτηριδιακής λοίμωξης σε ασθενείς με ΧΑΠ και ιδιαίτερα των λοιμογόνων Caga (+) στελεχών τα οποία ενοχοποιούνται για την παραγωγή προφλεγμονωδών κυτοκινών (όμοιες με αυτές που εμπλέκονται στην παθογένεια της ΧΑΠ) θεωρείται πιθανός ο παθογενετικός ρόλος του ελικοβακτηριδίου στη ΧΑΠ. Αντίθετα, περιορισμένος φαίνεται να είναι ο ρόλος τον οποίο διαδραματίζει το ελικοβακτηρίδιο στην εξέλιξη της νόσου, καθώς δε διαπιστώθηκε συσχέτιση της Η pylori οροθετικότητας και της CagA οροθετικότητας με τη βαρύτητα της ΧΑΠ Σίγουρα στο μέλλον απαιτούνται μελέτες σε μεγαλύτερο αριθμό ασθενών οι οποίες να ελέγχουν την ορθότητα των ευρημάτων μας.

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