Revision 6

#88005Store at +4C

 

()

Species Cross Reactivity

H

UniProt ID:

#P0DTC2

Entrez-Gene Id:

#43740568

Cell Signaling Technology

Orders: 877-616-CELL (2355) [email protected]

Support: 877-678-TECH (8324)

Web: [email protected] cellsignal.com

3 Trask LaneDanversMassachusetts01923USA
For Research Use Only. Not for Use in Diagnostic Procedures.
Product Includes Product # Quantity Color Storage Temp
Spike Multi-Domain Protein Coated Microwells 43037 96 tests +4C
Anti-Human IgG, HRP-linked Antibody (ELISA Formulated) 94210 1 ea Red (Lyophilized) +4C
Sample Diluent A 71637 25 ml +4C
HRP Diluent 13515 11 ml Red +4C
ELISA Wash Buffer (20X) 9801 25 ml +4C
TMB Substrate 7004 11 ml +4C
STOP Solution 7002 11 ml +4C
Sealing Tape 54503 2 ea +4C
ELISA Kit #88005 Positive Control 57510 1 ea +4C
ELISA Kit #88005 Negative Control 73072 1 ea +4C

*The microwell plate is supplied as 12 8-well modules - Each module is designed to break apart for 8 tests.

Description

The SARS-CoV-2 Spike Protein Multi-Domain (S1-NTD, RBD, S1, S2) Serological IgG ELISA Kit is a solid-phase ELISA that detects binding of human IgG to four domains of SARS-CoV-2 spike protein (S-protein): the S1-NTD, RBD, S1, and S2 domains, individually. The four spike protein domains have each been individually coated onto microwells (see plate map for location/color-coding), such that twenty-four tests are provided for each spike protein domain (96 tests total). After incubation with sample, the human IgG specific for each spike protein domain is captured on the plate. The wells are then washed to remove unbound material. Anti-Human IgG, HRP-linked antibody is then used to recognize the bound IgG. HRP substrate, TMB, is added to develop color. The magnitude of optical density for this developed color is proportional to the quantity of IgG specific for each spike protein domain.

*Antibodies in this kit are custom formulations specific to kit.

Specificity/Sensitivity

The SARS-CoV-2 Spike Protein Multi-Domain (S1-NTD, RBD, S1, S2) Serological IgG ELISA Kit detects endogenous levels of human IgG directed to the S1-NTD (16-316), RBD (318-541), S1 (16-681), and S2 (686-1208) domains of SARS-CoV-2 spike protein (S-protein).

Background

The cause of the COVID-19 pandemic is a novel and highly pathogenic coronavirus, termed SARS-CoV-2 (severe acute respiratory syndrome coronavirus-2). SARS-CoV-2 is a member of the Coronaviridae family of viruses (1). The genome of SARS-CoV-2 is similar to other coronaviruses, and is comprised of four key structural proteins: S, the spike protein, E, the envelope protein, M, the membrane protein, and N, the nucleocapsid protein (2). Coronavirus spike proteins are class I fusion proteins and harbor an ectodomain, a transmembrane domain, and an intracellular tail (3,4). The highly glycosylated ectodomain projects from the viral envelope surface and facilitates attachment and fusion with the host cell plasma membrane. The ectodomain can be further subdivided into host receptor-binding domain (RBD) (S1) and membrane-fusion (S2) subunits, which are produced upon proteolysis by host proteases at S1/S2 and S2’ sites. S1 and S2 subunits remain associated after cleavage and assemble into crown-like homotrimers (2,4). In humans, both SARS-CoV and SARS-CoV-2 spike proteins utilize the angiotensin-converting enzyme 2 (ACE2) protein as a receptor for cellular entry (5-7). Spike protein subunits represent a key antigenic feature of coronavirus virions, and therefore represent an important target of vaccines, novel therapeutic antibodies, and small-molecule inhibitors (8,9).

  1. Zhou, P. et al. (2020) Nature 579, 270-3.
  2. Tortorici, M.A. and Veesler, D. (2019) Adv Virus Res 105, 93-116.
  3. Li, F. et al. (2006) J Virol 80, 6794-800.
  4. Li, F. (2016) Annu Rev Virol 3, 237-61.
  5. Shang, J. et al. (2020) Nature 581, 221-4.
  6. Wrapp, D. et al. (2020) Science 367, 1260-3.
  7. Yan, R. et al. (2020) Science 367, 1444-8.
  8. Yuan, Y. et al. (2017) Nat Commun 8, 15092.
  9. Amanat, F. and Krammer, F. (2020) Immunity 52, 583-9.

Background References

    Cross-Reactivity Key

    H: human M: mouse R: rat Hm: hamster Mk: monkey Vir: virus Mi: mink C: chicken Dm: D. melanogaster X: Xenopus Z: zebrafish B: bovine Dg: dog Pg: pig Sc: S. cerevisiae Ce: C. elegans Hr: horse GP: Guinea Pig Rab: rabbit All: all species expected

    Trademarks and Patents

    Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.
    All other trademarks are the property of their respective owners. Visit cellsignal.com/trademarks for more information.

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    Orders: 877-616-CELL (2355)•[email protected]• Support: 877-678-TECH (8324)•[email protected]• Web: cellsignal.com
    For Research Use Only. Not for Use in Diagnostic Procedures.
    Revision 6
    #88005

    SARS-CoV-2 Spike Protein Multi-Domain (S1-NTD, RBD, S1, S2) Serological IgG ELISA Kit

    SARS-CoV-2 Spike Protein Multi-Domain (S1-NTD, RBD, S1, S2) Serological IgG ELISA Kit: Image 1 Expand Image
    Patient Testing: Patient samples were tested using the SARS-CoV-2 Spike Protein Multi-Domain (S1-NTD, RBD, S1, S2) Serological IgG ELISA Kit #88005. Serum or plasma was obtained from donors who had been diagnosed with SARS-CoV-2 (diagnosed positive n=59*) or from presumed uninfected donors collected prior to the SARS-CoV-2 outbreak (presumed negative n=86). Samples were heat-inactivated (56°C for 30 min) and diluted 1:400 prior to running the assay, as described in the protocol. Samples were considered positive, negative, or inconclusive based on the criteria described in the “Data Analysis” section of the protocol for the kit. Positive percent agreement (PPA) and negative percent agreement (NPA) were calculated from these data.

    *Positive reference samples were from patient donors with a positive SARS-CoV-2 diagnosis. However, a positive SARS-CoV-2 diagnosis will not always correlate with a positive response aganst each spike domain protein in this IgG Serological ELISA as differences in disease severity, timing of sample collection relative to disease onset, and patient profiles may affect presence and abundance of antibodies against each spike domain in the reference sample.

    Note: We are continuing to test more samples as available. For the most up-to-date set of data, always refer to the product page for #88005 on the website.

    SARS-CoV-2 Spike Protein Multi-Domain (S1-NTD, RBD, S1, S2) Serological IgG ELISA Kit: Image 2 Expand Image
    Intra-Assay Precision: Three different serum samples were each tested in 16 replicates using a single assay kit of SARS-CoV-2 Spike Protein Multi-Domain (S1-NTD, RBD, S1, S2) Serological IgG ELISA Kit #88005. Intra-assay CV (%) was calculated for each sample, and each replicate was correctly identified as being positive or negative when compared to the Negative Control using the cutoff criteria described in the attached protocol.
    SARS-CoV-2 Spike Protein Multi-Domain (S1-NTD, RBD, S1, S2) Serological IgG ELISA Kit: Image 3 Expand Image
    Inter-Assay Precision: Six different assay kits from one lot of material were tested using 3 different serum samples, Positive Control, and Negative Control run in duplicate wells. Inter-assay CV (%) was calculated for each sample, and each assay kit correctly identified the samples as being positive or negative when compared to the Negative Control using the cutoff criteria described in the attached protocol.

    *The included Positive Control in the kit is intended to give a positive signal for the RBD and S1 coated microwell strips, and is NOT a Positive Control for the S1-NTD or S2 microwells.

    SARS-CoV-2 Spike Protein Multi-Domain (S1-NTD, RBD, S1, S2) Serological IgG ELISA Kit: Image 4 Expand Image
    Plate map for SARS-CoV-2 Spike Protein Multi-Domain (S1-NTD, RBD, S1, S2) Serological IgG ELISA Kit #88005. Schematic representation of a 96-well plate depicting the color-code of the reagents used to detect binding of human IgG to four domains of SARS-CoV-2 spike protein (S-protein): the S1-NTD (blue, columns 1-3), RBD (orange, columns 4-6), S1 (gray, columns 7-9), and S2 (black, columns 10-12).
    SARS-CoV-2 Spike Protein Multi-Domain (S1-NTD, RBD, S1, S2) Serological IgG ELISA Kit: Image 5 Expand Image
    Spike Protein S1-NTD Serological IgG ELISA Patient Testing Results: Patient samples were tested using the SARS-CoV-2 Spike Protein Multi-Domain (S1-NTD, RBD, S1, S2) Serological IgG ELISA Kit #88005. Serum/plasma samples were heat-inactivated (56°C for 30 min) and diluted 1:400 prior to running the assay, as described in the protocol. Background subtracted absorbance values at 450 nm are plotted for each serum/plasma sample, with values corresponding to samples obtained from donors who had been diagnosed with SARS-CoV-2 (diagnosed positive n=59*) on the right, and values corresponding to samples from presumed uninfected donors collected prior to the SARS-CoV-2 outbreak (presumed negative n=86) on the left. Also shown is the cutoff line above which samples are considered positive (blue line), and the negative cutoff line (red line), below which samples are considered negative. Cutoff values were calculated as described in the “Data Analysis” section of the protocol for the kit.

    *Positive reference samples were from patient donors with a positive SARS-CoV-2 diagnosis. However, a positive SARS-CoV-2 diagnosis will not always correlate with a positive response against each spike domain protein in this IgG Serological ELISA as differences in disease severity, timing of sample collection relative to disease onset, and patient profiles may affect presence and abundance of antibodies against each spike domain in the reference sample.

    SARS-CoV-2 Spike Protein Multi-Domain (S1-NTD, RBD, S1, S2) Serological IgG ELISA Kit: Image 6 Expand Image
    Spike Protein RBD Serological IgG ELISA Patient Testing Results: Patient samples were tested using the SARS-CoV-2 Spike Protein Multi-Domain (S1-NTD, RBD, S1, S2) Serological IgG ELISA Kit #88005. Serum/plasma samples were heat-inactivated (56°C for 30 min) and diluted 1:400 prior to running the assay, as described in the protocol. Background subtracted absorbance values at 450 nm are plotted for each serum/plasma sample, with values corresponding to samples obtained from donors who had been diagnosed with SARS-CoV-2 (diagnosed positive n=59*) on the right, and values corresponding to samples from presumed uninfected donors collected prior to the SARS-CoV-2 outbreak (presumed negative n=86) on the left. Also shown is the cutoff line above which samples are considered positive (blue line), and the negative cutoff line (red line), below which samples are considered negative. Cutoff values were calculated as described in the “Data Analysis” section of the protocol for the kit.

    *Positive reference samples were from patient donors with a positive SARS-CoV-2 diagnosis. However, a positive SARS-CoV-2 diagnosis will not always correlate with a positive response against each spike domain protein in this IgG Serological ELISA as differences in disease severity, timing of sample collection relative to disease onset, and patient profiles may affect presence and abundance of antibodies against each spike domain in the reference sample.

    SARS-CoV-2 Spike Protein Multi-Domain (S1-NTD, RBD, S1, S2) Serological IgG ELISA Kit: Image 7 Expand Image
    Spike Protein S1 Domain Serological IgG ELISA Patient Testing Results: Patient samples were tested using the SARS-CoV-2 Spike Protein Multi-Domain (S1-NTD, RBD, S1, S2) Serological IgG ELISA Kit #88005. Serum/plasma samples were heat-inactivated (56°C for 30 min) and diluted 1:400 prior to running the assay, as described in the protocol. Background subtracted absorbance values at 450 nm are plotted for each serum/plasma sample, with values corresponding to samples obtained from donors who had been diagnosed with SARS-CoV-2 (diagnosed positive n=59*) on the right, and values corresponding to samples from presumed uninfected donors collected prior to the SARS-CoV-2 outbreak (presumed negative n=86) on the left. Also shown is the cutoff line above which samples are considered positive (blue line), and the negative cutoff line (red line), below which samples are considered negative. Cutoff values were calculated as described in the “Data Analysis” section of the protocol for the kit.

    *Positive reference samples were from patient donors with a positive SARS-CoV-2 diagnosis. However, a positive SARS-CoV-2 diagnosis will not always correlate with a positive response against each spike domain protein in this IgG Serological ELISA as differences in disease severity, timing of sample collection relative to disease onset, and patient profiles may affect presence and abundance of antibodies against each spike domain in the reference sample.

    SARS-CoV-2 Spike Protein Multi-Domain (S1-NTD, RBD, S1, S2) Serological IgG ELISA Kit: Image 8 Expand Image
    Spike Protein S2 Domain Serological IgG ELISA Patient Testing Results: Patient samples were tested using the SARS-CoV-2 Spike Protein Multi-Domain (S1-NTD, RBD, S1, S2) Serological IgG ELISA Kit #88005. Serum/plasma samples were heat-inactivated (56°C for 30 min) and diluted 1:400 prior to running the assay, as described in the protocol. Background subtracted absorbance values at 450 nm are plotted for each serum/plasma sample, with values corresponding to samples obtained from donors who had been diagnosed with SARS-CoV-2 (diagnosed positive n=59*) on the right, and values corresponding to samples from presumed uninfected donors collected prior to the SARS-CoV-2 outbreak (presumed negative n=86) on the left. Also shown is the cutoff line above which samples are considered positive (blue line), and the negative cutoff line (red line), below which samples are considered negative. Cutoff values were calculated as described in the “Data Analysis” section of the protocol for the kit.

    *Positive reference samples were from patient donors with a positive SARS-CoV-2 diagnosis. However, a positive SARS-CoV-2 diagnosis will not always correlate with a positive response against each spike domain protein in this IgG Serological ELISA as differences in disease severity, timing of sample collection relative to disease onset, and patient profiles may affect presence and abundance of antibodies against each spike domain in the reference sample.

    SARS-CoV-2 Spike Protein Multi-Domain (S1-NTD, RBD, S1, S2) Serological IgG ELISA Kit: Image 9 Expand Image
    Heatmap depicting SARS-CoV-2 Spike Protein Multi-Domain (S1-NTD, RBD, S1, S2) Serological IgG ELISA Kit #88005 patient testing results. Shown are the results for a subset of the patient serum/plasma samples that were tested for the presence of IgG antibodies against S1-NTD, RBD, S1, and/or S2 using kit #88005. Absorbance values (OD450 Value) corresponding to wells containing one of the four SARS-CoV-2 spike protein domains were used to generate the heatmap. Serum/plasma samples from a subset of donors who had been diagnosed with SARS-CoV-2 (diagnosed positive*) and from presumed uninfected donors collected prior to the SARS-CoV-2 outbreak (presumed negative) were heat-inactivated (56°C for 30 min) and diluted 1:400 prior to running the assay, as described in the protocol.

    NOTE: Absorbance 450 nm values should not be directly compared across different spike domain proteins, as they are not absolute values.

    *Positive reference samples were from patient donors with a positive SARS-CoV-2 diagnosis. However, a positive SARS-CoV-2 diagnosis will not always correlate with a positive response against each spike domain protein in this IgG Serological ELISA as differences in disease severity, timing of sample collection relative to disease onset, and patient profiles may affect presence and abundance of antibodies against each spike domain in the reference sample.

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    Orders: 877-616-CELL (2355)•[email protected]• Support: 877-678-TECH (8324)•[email protected]• Web: cellsignal.com
    For Research Use Only. Not for Use in Diagnostic Procedures.