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PF-07302048 : Qualification Report for a Single -plex Direct Luminex Ass ay (dLIA) for Quantitation of IgG
Antibodies to SARS -CoV -2 S1 Protein in Hum an Sera
VR-MQR -10211 , Ver. 2.0
PFIZER CONFIDENTIAL
Page 1Title: Qualification Report for a Single -plex Direct L uminex Ass ay (dLIA) for Quantitation
of IgG Antibodies to SARS- CoV -2 S1 Protein in Human Sera
Study Number: N/A
Parent Compound Number(s): PF-07302048
Alternative Compound Identifiers: N/A
Pfizer Vaccine Research and Development
401 N. Middletown Rd.
Pearl River, NY
090177e194a89aaf\Approved\Approved On: 18-Aug-2020 23:11 (GMT)
FDA-CBER-2021-5683-0779207
PF-07302048 : Qualification Report for a Single -plex Direct Luminex Ass ay (dLIA) for Quantitation of IgG
Antibodies to SARS -CoV -2 S1 Protein in Hum an Sera
VR-MQR -10211 , Ver. 2.0
PFIZER CONFIDENTIAL
Page 2Title: Qualification Report for a Single -plex Direct L uminex Ass ay (dLIA) for Quantitation
of IgG Antibodies to SARS -CoV -2 S1 Protein in Human Sera
PRINCIPAL INVESTIGATOR: Guy Singh , Principal Sc ientist
CONTRIBUTING SCIENTIST (S):Darryl Charles, Estee Dilli , Nathaniel Garcia
PREPARED BY:
Guy Singh
Principal Scientist, Clinical and Diagnostic Assay Development
APPROVED BY:
Danka Pavliakova
Senior Principal Scientist, Clinical and Diagnostic Assay Development
Warren Kalina
Director, Clinical and Diagnostic Assay Development
Charles Tan
Senior Director, Earl y Clinical Development Biostatistics
David Cooper
Executive Director, High -throughput Clinical Immunoassay s & Diagnostics
Michael Pride
Executive Director, Clinical and Diagnostic Assay Development
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FDA-CBER-2021-5683-0779208
PF-07302048 : Qualification Report for a Single -plex Direct Luminex Ass ay (dLIA) for Quantitation of IgG
Antibodies to SARS -CoV -2 S1 Protein in Hum an Sera
VR-MQR -10211 , Ver. 2.0
PFIZER CONFIDENTIAL
Page 3Title: Qualification Report for a Single -plex Direct L uminex Ass ay (dLIA) for Quantitation
of IgG Antibodies to SARS- CoV -2 S1 in Protein Human Sera
SYNOPSIS
This report documents the results of the statistical anal yses from assay qualification
experiments of a single -plex direct L uminex immunoassay (dLIA) for q uantitation of human
IgG antibodies to subunit 1 (S1) of the spike (S) protein of severe acute respiratory disease
coronavirus 2 (SARS -CoV -2)in human sera . The performance parameters examined in the
assay qualification were linearity , precision and standard curve bias . From these
data, the assay range, lower limit of quantitation, and descriptive statistics of assay
parameters were determined. Key qualification outcomes are highlighted in the table below.
Key Qualification Outcomes
Antigen Assay Range
Lower Bound Plate
Well IgG
Concentrat ion
(U/mL)aAssay Range
Upper Bound Plate
Well IgG
Concentration
(U/mL)aLLOQbSample
Dilution Adjusted
IgGConcentration
(U/mL)aAssay Precision
c
COV S1d0.002533 0.128 000 1.2665
a.Units/mL .
b.Lower Limit of Quantitation .
c.
d.Subunit 1 of the Spike protein of severe acute respiratory disease coronavirus 2 .
The data provided in this report support the qualification of the single -plex dLIA for
quantitation of human IgG antibodies that bind to the S1 protein of SARS- CoV -2 and
confirm tha t the assay is suitable for its intended use when performed in accordance w ith
standard operating procedures by qualified personnel.
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FDA-CBER-2021-5683-0779209
PF-07302048 : Qualification Report for a Single -plex Direct Luminex Ass ay (dLIA) for Quantitation of IgG
Antibodies to SARS -CoV -2 S1 Protein in Hum an Sera
VR-MQR -10211 , Ver. 2.0
PFIZER CONFIDENTIAL
Page 4TABLE OF CONTENTS
SYNOPSI S................................ ................................ ................................ ................................ .3
LIST OF TABLES ................................ ................................ ................................ ..................... 5
LIST OF FIGURES ................................ ................................ ................................ ................... 6
1. OBJECTI VES ................................ ................................ ................................ ........................ 7
2. INTRODUCTION ................................ ................................ ................................ ................. 7
3. GLOSSARY ................................ ................................ ................................ .......................... 8
4. EXPERI MENTAL OUTL INE................................ ................................ ............................... 9
4.1. Mater ials and Methods ................................ ................................ .............................. 9
4.1.1. Methods ................................ ................................ ................................ ........ 9
4.1.2. Critical Reagents ................................ ................................ ........................... 9
4.2. Experimental Design ................................ ................................ ............................... 10
4.2.1. Linearity ................................ ................................ .................... 10
4.2.2. Precision ................................ ................................ ................................ .....11
4.2.3. Standard Curve Bias ................................ ................................ ................... 11
5. STATI STICAL METHO DS................................ ................................ ................................ 11
5.1. Sampl e Results ................................ ................................ ................................ ........ 11
5.2. Statistical Analy ses................................ ................................ ................................ .11
5.2.1. Linearity ................................ ................................ .................... 12
5.2.2. Precision ................................ ................................ ................................ .....12
5.2.3. Standard Curve Bias ................................ ................................ ................... 13
5.2.4. Assay Range ................................ ................................ ............................... 13
5.2.5. Assay Precision (Intermediate Precision) ................................ ................... 14
5.2.6. Sample Quantitation Range ................................ ................................ ........ 14
5.2.6.1. L ower Limit of Quantitation ................................ ..................... 14
5.2.6.2. Upper Limit of Quantitation ................................ ...................... 15
5.2.7. Assay Run Performance ................................ ................................ ............. 15
5.2.7.1. ................................ ................................ .15
5.2.7.2. Standard Curve Parameters ................................ ....................... 15
5.2.7.3. Quality Control Samples ................................ ........................... 15
6. RESUL TS AND DI SCU SSION ................................ ................................ .......................... 16
6.1. L inearity Evaluation ................................ ................................ ............... 16
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FDA-CBER-2021-5683-0779210
PF-07302048 : Qualification Report for a Single -plex Direct Luminex Ass ay (dLIA) for Quantitation of IgG
Antibodies to SARS -CoV -2 S1 Protein in Hum an Sera
VR-MQR -10211 , Ver. 2.0
PFIZER CONFIDENTIAL
Page 56.2. Precision Evaluation ................................ ................................ ................................ 17
6.3. Standard Curve Bias Evaluation ................................ ................................ ............. 18
6.4. Assay Range Based on L inearity , Precision, and Standard Curve
Bias ................................ ................................ ................................ ............................ 19
6.5. Assay Precision (Intermediate Precision) ................................ ............................... 19
6.6. Sample Quantitation Range ................................ ................................ ..................... 20
6.6.1. L ower Limit of Quantitation ................................ ................................ .......20
6.6.2. Upper Limit of Quantitation ................................ ................................ .......20
6.7. Assay Run Performance ................................ ................................ .......................... 20
6.7.1. ................................ ................................ .................. 20
6.7.2. Standard Curves ................................ ................................ .......................... 21
6.7.3. Quality Control Samples.............................................................................21
7. CONCLUSION ................................ ................................ ................................ .................... 22
8. DEVIATIONS ................................ ................................ ................................ ..................... 22
9. SUPPORTING DOCUME NTATION ................................ ................................ ................. 22
10. SUPPL EMENTAL INF ORMATION ................................ ................................ ............... 22
11. SUPPORTIVE FIGUR ES................................ ................................ ................................ ..24
12. SUPPORTIVE TABLE S ................................ ................................ ................................ ...25
LIST OF TABLES
Table 1. Terms and Definitions ................................ ................................ ................ 8
Table 2. SOPs and Robotic Methods within Scope of this Protocol ........................ 9
Table 3. List of Critical Reagents ................................ ................................ ........... 10
Table 4. Well Concentration Range Based on L inearit y Data .............. 17
Table 5. Well Concentration Range Based on Precision Data ............................... 18
Table 6. Well Concentration Range Based on Reference Standard Curve
Bias ................................ ................................ ................................ ........... 19
Table 7. Final IgG Assay Range – Well Concentration (Units/mL) ...................... 19
Table 8. .......... 19
Table 9. Lower Limit of Quantitation (LLOQ) ................................ ...................... 20
Table 10. ................................ ................................ ................ 21
Table 11. Descriptive Statistics for Standard Curve Parameters.............................. 21
Table 12. Descriptive Statistics for QCS ................................ ................................ ..22
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FDA-CBER-2021-5683-0779211
PF-07302048 : Qualification Report for a Single -plex Direct Luminex Ass ay (dLIA) for Quantitation of IgG
Antibodies to SARS -CoV -2 S1 Protein in Hum an Sera
VR-MQR -10211 , Ver. 2.0
PFIZER CONFIDENTIAL
Page 6LIST OF FIGURES
Figure 1. Linearity Plot for S1 IgG dLIA ................................ ............... 16
Figure 2. Precision Plot for S1 IgG dLIA ................................ ................................ 17
Figure 3. Standard Curve Bias Plot for S1 IgG dLIA ................................ .............. 18
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FDA-CBER-2021-5683-0779212
PF-07302048 : Qualification Report for a Single -plex Direct Luminex Ass ay (dLIA) for Quantitation of IgG
Antibodies to SARS -CoV -2 S1 Protein in Hum an Sera
VR-MQR -10211 , Ver. 2.0
PFIZER CONFIDENTIAL
Page 7Title: Qualification Report for a Single -plex Direct L uminex Ass ay (dLIA) for Quantitation
of IgG Antibodies to SARS- CoV -2 S1 Protein in Human Sera
Study Number: NA
Functional Area: Vaccine Research and Development
Test Facility : Pfizer Vaccine Research, 401 North Middletown Road,
Pearl River, NY 10965
Study Initiation Date: 15Apr2020
Study Completion Date: 01Jul2020
1.OBJECTIVES
This report summarizes the methodology , results, and sta tistical anal ysis of the assay
qualification for a single -plex direct d LIA for q uantitation of IgG antibodies to the S1 protein
of SARS -CoV -2 in human serum , described in VR-TM-10293 (see Section 9 ). Assay
qualification provides documented evidence that the assay , when performed in accordance
with standard operating procedures (SOPs) b y qualified personnel, is suitable for the
intended to quantify S1-specific IgG antibody concentrations in human sera.
2.INTRODUCTION
The single -plex SARS -CoV -2 S1 IgG dLIA measures specific IgG antibodies binding to the
S1 domain of the SARS -CoV -2 spike protein (S). This assay is described in the test method,
VR-TM-10293 (see Section 9) and is based on the Luminex MagPlex xMAP technology .
Magnetic, fluorescent Luminex microspheres are coated with according to
VR-SOP-LC-112951andthen S1 protein. S1 coated microspheres
are added to serum samples, diluted in assay buffer and incubated, with shaking, in a 96- well
microtiter assay plates for hours at °C. Unbound assay components are removed b y
washing and a purified goat anti- human IgG secondary antibody
is added to the reaction wells. The secondary antibody is incubated for minutes at
°C. Unbound assay components are removed by washing and the reaction is read on
a Bio -Plex reader. The fluorescent protein coupled to the secondary antibody allows
measurement of the specific antibody bound to the antigen coated microspheres.
Fluorescence i s expressed as median fluorescent intensities (MFI), and the assay results are
calculated against a reference standard with arbitrary assigned concentration of
100.00 Units/mL . All sera are testedin duplicate from independentl y generated samples .
Test samples are initiall y tested at dilutions,
dilutions are used to increase the likelihood that at least one result for an y sample will fall
within the useable range of the standard curve. Using acceptance crit eriaestablished in this
report , failed plates and samples may be repeated as required.
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FDA-CBER-2021-5683-0779213
PF-07302048 : Qualification Report for a Single -plex Direct Luminex Ass ay (dLIA) for Quantitation of IgG
Antibodies to SARS -CoV -2 S1 Protein in Hum an Sera
VR-MQR -10211 , Ver. 2.0
PFIZER CONFIDENTIAL
Page 8The final assay results are expressed as the geometric mean concentration (GMC) of all
sample dilutions that produced a valid assay result within the assay range. GM C results that
are below the Lower Limit of Quantitation (LLOQ) are reported as Below Limit of
Quantitation (BLQ).
3.GLOSSARY
Table 1.Terms and Definitions
Term Definition
Assay Range Range of antibody concentrations that can be measured in the assay
plate w ell that have linearity, precision, and standard curve bias within the limits
described below . The assay range is generated from the most conservative lower and
upper w ell concentration limits based on linearity ,precision , and standard curve
bias.
Beads Luminex Microspheres
BLQ Below Limit of Quantitation
CDAD Clinical and Diagnostic Assay Development
COV ID-19 Coronavirus Disease 2019
dLIA Direct Luminex ImmunoAssay
GMC Geom etric Mean Concentration
HCID High -throughput Clinical Immunoassays & Diagnostics
IgG Immunoglobulin G
LIMS Laboratory Information Management System
LLOQ Lower Limit of Quantitation -the lowest dilution -adjusted sample concentration that can
be determined with precision and falls within the linear portion of the
assay range .
MFI Median Fluorescent Intensity
mL Milliliter
NHP Non-human prim ate
QA Quality Assurance
QCS Quality Control Sample
PE Phycoery thrin
RT Room Tem perature 18-25°C
S Spike glycoprotein of severe acute respiratory disease coronavirus 2
S1 Subunit 1 of the Spike protein
SARS -CoV-2 Severe acute respiratory disease coronavirus 2
Sample
Concentrationantibody concentration calculated as the from the
sample that fall within the assay range and the lower and upper parameter limits
of the standard curve that is run on the same assay plate.
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FDA-CBER-2021-5683-0779214
PF-07302048 : Qualification Report for a Single -plex Direct Luminex Ass ay (dLIA) for Quantitation of IgG
Antibodies to SARS -CoV -2 S1 Protein in Hum an Sera
VR-MQR -10211 , Ver. 2.0
PFIZER CONFIDENTIAL
Page 9Table 1.Terms and Definitions
Term Definition
Sample Range Range of sample concentrations that can be quantified without pre -
dilution. The sample range is bounded by the LLOQ and ULOQ. Results below LLOQ
are reported as BLQ.
SOP Standard Operating Procedure
TM Test Method
ULOQ Upper Limit of Quantitation -the highest sample concentration that can
be determined with precision and falls within the linear portion of the
assay range
VRD Vaccine Research and Development
Well
concentration antibody concentrations in the assay plate w ells
4.EXPERIMENTAL OUTLINE
4.1.Materials and Methods
Unique reagents prepared for this assessment are listed below, and all other materials are
described in the SOPs referenced in Table 2. Spe cific details regarding reagent catalog and
lot numbers and expiration, as well as the instrument identification numbers and maintenance
details w eredocumented in the assay worksheets within data packages.
4.1.1. Methods
Table 2. SOPs and Robotic Methods within Scope of this Protocol
SOP Number Work Described in SOP Robotic Method Nam e
VR-TM-10293
(see Section 9)Test method N/A
VR-SOP-LC-112951Bead coating method N/A
VR-SOP-LC-106272Preparation of buffers N/A
VR-SOP-LC-111863Sample preparation using robot
4.1.2. Critical Reagents
The critical reagents, including antigen -coated microspheres (beads), reference standard,
QCS and secondary antibodies used in this assay qualification are listed in
Table 3.
The reference standard serum is a human sera (COVID-19
sera drawn at least day s after PCR -confirmed diagnosis from patients
years of age) and ass igned an arbitrary concentration of 100.00 U/mL of IgG antibodies to
the S1 antigen.
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FDA-CBER-2021-5683-0779215
PF-07302048 : Qualification Report for a Single -plex Direct Luminex Ass ay (dLIA) for Quantitation of IgG
Antibodies to SARS -CoV -2 S1 Protein in Hum an Sera
VR-MQR -10211 , Ver. 2.0
PFIZER CONFIDENTIAL
Page 10Table 3.List of Critical Reagents
S1 coated microspheres
Secondary antibody
Reference Standard Serum
COVID19 IgG STDa PCR confirmed
Covid -19 sera
QCS1
COVID19 IgG QC1b
QCS2
COVID19 IgG QC2c
QCS3
COVID19 IgG QC3d
a.Assay Standard .
b.Assay Quality control serum 1 .
c.Assay Quality control serum 2 .
d.Assay Quality control serum 3 .
4.2.Experimental Design
The following sections describe the experimental designs for evaluating the
linearity , precision and standard curves.
4.2.1. Linearity
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FDA-CBER-2021-5683-0779216
PF-07302048 : Qualification Report for a Single -plex Direct Luminex Ass ay (dLIA) for Quantitation of IgG
Antibodies to SARS -CoV -2 S1 Protein in Hum an Sera
VR-MQR -10211 , Ver. 2.0
PFIZER CONFIDENTIAL
Page 114.2.2. Precision
Precision describes the closeness of measurements for a sample tested multiple times.
Precision is a measure of assay variabilit y that contains both repeatability and intermediate
precision. Repeatability measures the assay variability , usuall y within one assay run,
wherea s intermediate precision measures the variability within the same laboratory while
taking into account relevant sources of variation due to operating conditions ( eg,different
analysts, time, reagent lots).
The precision of the assay was examined using a panel of COVID -19
serawith IgG antibody concentrations intended to span the expected assay range and
pre-pandemic human serum samples that have low or no specific antibodies for a total
of samples. Refer to Supportive Table 12.2 for the precision samples used.
All samples were tested in the assay at dilutions performed b y
a single workstation
, using the methods developed for clinical testing as
described in VR -SOP-LC-111863. Refer to Supportive Figure 11.2 for the assay plate
layout used for the precision experiments.
Preci sion measurements of the samples , described above, were performed following
VR-TM-10293 (see Section 9)over
Results were anal yzed as described in Section 5.2.2 and
Section 5.2.5 .
4.2.3. Standard Curve Bias
5. STATISTICAL METHODS
5.1.Sample Results
5.2. Statistical Analyses
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PF-07302048 : Qualification Report for a Single -plex Direct Luminex Ass ay (dLIA) for Quantitation of IgG
Antibodies to SARS -CoV -2 S1 Protein in Hum an Sera
VR-MQR -10211 , Ver. 2.0
PFIZER CONFIDENTIAL
Page 125.2.1. Linearity
5.2.2. Precision
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PF-07302048 : Qualification Report for a Single -plex Direct Luminex Ass ay (dLIA) for Quantitation of IgG
Antibodies to SARS -CoV -2 S1 Protein in Hum an Sera
VR-MQR -10211 , Ver. 2.0
PFIZER CONFIDENTIAL
Page 135.2.3. Standard Curve Bias
5.2.4. Assay Range
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PF-07302048 : Qualification Report for a Single -plex Direct Luminex Ass ay (dLIA) for Quantitation of IgG
Antibodies to SARS -CoV -2 S1 Protein in Hum an Sera
VR-MQR -10211 , Ver. 2.0
PFIZER CONFIDENTIAL
Page 145.2.5. Assay Precision (Intermediate Precision)
5.2.6. Sample Quantitation Range
The quantitation limits are defined as the sample concentration s that are between the LLOQ
and the ULOQ.
5.2.6.1. Lower Limit of Quantitation
The lower limit of quantitation (LLOQ) is the lowest sample concentration in the assay that
can be determined with precision and falls within the lin ear portion of the
assay range .
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PF-07302048 : Qualification Report for a Single -plex Direct Luminex Ass ay (dLIA) for Quantitation of IgG
Antibodies to SARS -CoV -2 S1 Protein in Hum an Sera
VR-MQR -10211 , Ver. 2.0
PFIZER CONFIDENTIAL
Page 155.2.6.2. Upper Limit of Quantitation
The upper limit of quantitation (ULOQ) is the highest sample concentration that can be
determined with precision and falls into the linear assay range .
5.2.7. Assay Run Performance
5.2.7.1.
5.2.7.2. Standard Curve Parameters
5.2.7.3. Quality Control Samples
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PF-07302048 : Qualification Report for a Single -plex Direct Luminex Ass ay (dLIA) for Quantitation of IgG
Antibodies to SARS -CoV -2 S1 Protein in Hum an Sera
VR-MQR -10211 , Ver. 2.0
PFIZER CONFIDENTIAL
Page 166. RESULTS AND DISCUSSI ON
Qualification data used for the results of the anal yses described herein are listed in
VR-MQR -10211-ATT01 (see Section 9).
6.1. Linearity Evaluation
Figure 1. Linearity Plot for S1IgG dLIA
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PF-07302048 : Qualification Report for a Single -plex Direct Luminex Ass ay (dLIA) for Quantitation of IgG
Antibodies to SARS -CoV -2 S1 Protein in Hum an Sera
VR-MQR -10211 , Ver. 2.0
PFIZER CONFIDENTIAL
Page 17Table 4.Well Concentration Range Based on Linearity Data
6.2.Precision Evaluation
Figure 2.Precision Plot for S1IgG dLIA
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PF-07302048 : Qualification Report for a Single -plex Direct Luminex Ass ay (dLIA) for Quantitation of IgG
Antibodies to SARS -CoV -2 S1 Protein in Hum an Sera
VR-MQR -10211 , Ver. 2.0
PFIZER CONFIDENTIAL
Page 18Table 5.Well Concentration Range Based on Precision Data
6.3.Standard Curve Bias Evaluation
Figure 3. Standard Curve Bias Plot for S1IgG dLIA
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FDA-CBER-2021-5683-0779224
PF-07302048 : Qualification Report for a Single -plex Direct Luminex Ass ay (dLIA) for Quantitation of IgG
Antibodies to SARS -CoV -2 S1 Protein in Hum an Sera
VR-MQR -10211 , Ver. 2.0
PFIZER CONFIDENTIAL
Page 19Table 6.Well Concentration Range Based on Reference Standard Curve Bias
6.4.Assay Range Based on Linearity, Precision, and Standard Curve Bias
Table 7.Final Ig GAssay Range –Well Concentration (U nits/mL)
Antigen
Linearity
RangePrecision
RangeStandard Curve
RangeFinal Assay Range
Lower Upper
COV S1a0.002533 0.128000
a. S ubunit 1 of the Spike protein of severe acute respiratory disease coronavirus 2 .
6.5.Assay Precision (Intermediate Precision)
The intermediate precision of the assay was evaluated using the as described in
Section 5.2.5 and the results are summarized in Table 8.
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FDA-CBER-2021-5683-0779225
PF-07302048 : Qualification Report for a Single -plex Direct Luminex Ass ay (dLIA) for Quantitation of IgG
Antibodies to SARS -CoV -2 S1 Protein in Hum an Sera
VR-MQR -10211 , Ver. 2.0
PFIZER CONFIDENTIAL
Page 206.6.Sample Quantitation Range
6.6.1. Lower Limit of Quantitation
The LLOQ is the lowest sample IgG concentration in the assay that is precise
and falls into the linear assay range. Unlike the lower limit of the assay
range that is defined as a well concentration, the LLOQ is a dilution -adjusted sample
concentra tion. As described in Section 5.2.6.1
, LLOQ is defined as the dilution -adjusted
lower l imit of the assay range. Table 9 lists the LLOQ value for the SARS CoV -2 S1 IgG
dLIA.
Table 9.Lower Limit of Quantitation (LLOQ)
Antigen LLOQ (U nits/mL)
COV S1a1.2665
a.Subunit 1 of the Spike protein of severe acute respiratory disease coronavirus 2 .
6.6.2. Upper Limit of Quantitation
6.7.Assay Run Performance
6.7.1.
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PF-07302048 : Qualification Report for a Single -plex Direct Luminex Ass ay (dLIA) for Quantitation of IgG
Antibodies to SARS -CoV -2 S1 Protein in Hum an Sera
VR-MQR -10211 , Ver. 2.0
PFIZER CONFIDENTIAL
Page 21Table 10.
6.7.2. Standard Curve s
Descriptive statistics for the reference standard curves for S1IgGare listed in Table 11.
Table 11presents suitability limits on standard curves which were calculated as described in
Section 5.2.7.2 .
Table 11. Descriptive Statistics for Standard Curve Pa rameter s
6.7.3. Quality Control Samples
Descriptive statistics for the QCS samples from the qualification data for the SARS -CoV -2
S1 IgG dLIA are listed in Table 12.
Table 12also shows QCS limits on the plates (refer to
Section 5.2.7.3 ).
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PF-07302048 : Qualification Report for a Single -plex Direct Luminex Ass ay (dLIA) for Quantitation of IgG
Antibodies to SARS -CoV -2 S1 Protein in Hum an Sera
VR-MQR -10211 , Ver. 2.0
PFIZER CONFIDENTIAL
Page 22Table 12. Descriptive Statistics for QCS
7.CONCLUSION
The data provided in this report support the qualification of the single -plex SARS -CoV -2 S1
IgGdLIA for quantitating S1 specific IgGinhuman sera . The assay is suitable for the
intended use when performed in accordance with standard operating procedures by qualified
personnel. The assay limits established from the data generated in this qualification will be
used to support assay suitability during clinical testing until such limits are refined in a future
validation .
8.DEVIATIONS
NA
9. SUPPORTING DOCUMENTA TION
1.VR-TM-10293, Single -plex L uminex Assay for Quantitation of IgG Antibodies to
SARS -CoV-2 S1 protein in Human Serum.
2.VR-MQR -10211 -ATT01, Supportive Data for VR- MQR -10211 .
3.VR-SOP-LC-11120, Data Review Procedures for Direct Luminex I mmunoassay s in
LIMS v6 .
10. SUPPLEMENTAL INFORMA TION
1. VR-SOP-LC-11295, Preparation and Evaluation of Coated Microspheres
for use in Direct Luminex Assay s.
2. VR-SOP-LC-10627 , Preparation of Assay Buffers and Solution.
3. VR-SOP-LC-11186 , Standard Operating Procedure for Running Method
Using Microlab STAR Robot .
4.
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PF-07302048 : Qualification Report for a Single -plex Direct Luminex Ass ay (dLIA) for Quantitation of IgG
Antibodies to SARS -CoV -2 S1 Protein in Hum an Sera
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PFIZER CONFIDENTIAL
Page 235. VR-DTN -11380, Planned Deviation from VR- SOP-FE-10111 Bio- Plex Operation
and Maintenance .
6. VR-SOP-FE-10111, BioPlex System Operation and Maintenance .
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PF-07302048 : Qualification Report for a Single -plex Direct Luminex Ass ay (dLIA) for Quantitation of IgG
Antibodies to SARS -CoV -2 S1 Protein in Hum an Sera
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PFIZER CONFIDENTIAL
Page 2411.SUPPORTIVE FIGURES
11.1. Plate Layout for Linearity Experiments ..........................................24
11.2. SARS-C oV-2 S1 IgG dLIA Assay Plate Layout for Routine Testing............................24
11.1. Plate Layout for Linearity Experiments
11.2. SARS -CoV -2 S1 IgG dLIA Assay Plate Layout for Routine Testing
The assay plate lay out in Supportive Figure 11.2 is used for routine testing.
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PF-07302048 : Qualification Report for a Single -plex Direct Luminex Ass ay (dLIA) for Quantitation of IgG
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PFIZER CONFIDENTIAL
Page 2512.SUPPORTIVE TABLES
12.1. Linearity Samples ..........................................................................................25
12.2. Sample Panel for Precision Evaluation ................................ ................................ ........... 26
12.3. Precision of COVS1 Well Titers Sample................................ ....26
12.4. ................................ .................. 28
12.1. Linearity Samples
Sample Num ber Sample ID
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PF-07302048 : Qualification Report for a Single -plex Direct Luminex Ass ay (dLIA) for Quantitation of IgG Antibodies to SARS -CoV -2 S1 Protein in Human
Sera
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PFIZER CONFIDENTIAL
Page 2612.2. Sample Panel for Precision Evaluation
Sample Number ID Sample Number ID Sample Number ID Sample Number ID
12.3. Precision of COVS1 Well Titers Sample
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PF-07302048 : Qualification Report for a Single -plex Direct Luminex Ass ay (dLIA) for Quantitation of IgG Antibodies to SARS -CoV -2 S1 Protein in Human
Sera
VR-MQR -10211 , Ver. 2.0
PFIZER CONFIDENTIAL
Page 2712.3. Precision of COVS1 Well Titers Sample
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PF-07302048 : Qualification Report for a Single -plex Direct Luminex Ass ay (dLIA) for Quantitation of IgG
Antibodies to SARS -CoV -2 S1 Protein in Hum an Sera
VR-MQR -10211 , Ver. 2.0
PFIZER CONFIDENTIAL
Page 28
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